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2012.igem.org - User contributions [en]
2024-03-29T00:26:28Z
From 2012.igem.org
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http://2012.igem.org/File:120922.PNG
File:120922.PNG
2012-10-27T04:04:58Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120921-2.PNG
File:120921-2.PNG
2012-10-27T04:04:49Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120921.PNG
File:120921.PNG
2012-10-27T04:04:44Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120920.PNG
File:120920.PNG
2012-10-27T04:03:37Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120919-3.PNG
File:120919-3.PNG
2012-10-27T04:03:31Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T03:59:29Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
{{:Team:KAIST_Korea/navi}}<br />
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<br />
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font-family: 'Open Sans', sans-serif;<br />
color:#848383;<br />
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font-size:14pt;<br />
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#kaistcontent section<br />
{<br />
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border-width:1px;<br />
border-color:#EEE;<br />
padding:2px 10px 10px 10px;<br />
margin:0px 0px 35px 0px;<br />
background-image:url('https://static.igem.org/mediawiki/2012/a/a0/KAIST_labnote_logo.png');<br />
background-repeat:no-repeat;<br />
opacity:0.9;<br />
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color:#f4f4f4;<br />
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</style><br />
</head><br />
<br />
<body><br />
<div id="main"><br />
<img id="top-img" src="https://static.igem.org/mediawiki/2012/6/6d/Back_home.PNG"/><br />
<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
</div><br />
<br />
<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/5e/120903.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no band for 1516 gene. Band of all three genes appeared at the right size.</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></br><br />
We, </br><br />
1. changed the fraction of primers down to 0.5ul each, (lane 1)</br><br />
2. changed the polymerase to pfu-x, (lane3)</br><br />
3. used hybrid primers ( TOPO forward, Gibson reverse : lane 4 and TOPO reverse, Gibson forward : lane 5)</br><br />
Lane 2 was the control experiment with Hotstartaq polymerase.</br><br />
</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of piBR181 vector containing cells. </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 3ml pre-culture on five 10ml tubes for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer. </br></br></span><br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/ba/120905_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragment, we did PCR amplification of each gene once more. Correct size of bands appeared. </br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1197</i> <i>1516</i> PCR amplification </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/94/120905-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1197 once more. And we did PCR amplification of 1516 gene once more with the same primers. Correct size of bands appeared for 1197 gene, but there was no band for 1516 gene. . </br></br></span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1198</i> <i>1516</i>PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8c/120906.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1198 once more. And we did PCR amplification of 1516 gene once more with the same primers. For 1516 gene, we couldn’t have right size of band for new primers. So we decided to use the fragments of 1516 that does not contain terminator sequence. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">Pre-culture of <i>0109</i>, <i>1191</i>, FDH containing cells.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">We did 5ml pre-culture on 10ml tube for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer.<br />
<br />
</br></br></span><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">FDH pTrc & pBAD/<i>0109</i> pBAD/<i>1191</i> pBAD MG1655 expression check (sampling and induction)</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Procedure</b></br></br><br />
<span id="little">We seeded cells (500λ of cells put into 50mL LB with 50λ Ampicillin) and checked Optical Density. According to the O.D. value, we induced 37℃ and 30℃ sample and did sampling. Each samples were induced by different conditions ; 1mM and 2mM of IPTG, 10mM and 30mM of arabinose.</br></br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">Moth_0109 expression optimization _ gel picture</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/64/120909.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 59.94kDa protein, formate--tetrahydrofolate ligase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of 0109 gene on pBAD vector has failed and decided to try once more.</br></br></span><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Moth_1191 expression optimization _ gel picture </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/e/e0/120909_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 33.92kDa protein, methylenetetrahydrofolate reductase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of 1191 gene on pBAD vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc expression optimization _ gel picture </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/bf/120909-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 40.37kDa protein, Formate dehydrogenase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of FDH gene on pTrc vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pBAD expression optimization _ gel picture</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/6d/120909-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 40.37kDa protein, Formate dehydrogenase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of FDH gene on pTrc vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Primer design to PCR-amplify piBR181 vector.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d5/1209094.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For the Gibson assembly with RE digestion of vector, we had several problems with de-phosphorylation and purification after de-phosphorylation. Therefore we decided to PCR-amplify the piBR181 vector and use the PCR product in Gibson assembly. </br></br></span><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">PCR amplification of 1202 gene with new primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/ad/120910.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">The correct size of band appeared for the second PCR product, but there was no band appeared for the first PCR product. </br></br></span><br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">PCR amplification of piBR181 vector</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/66/120911.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">The correct size of piBR181 vector pcr product is 5297bp. There appeared the right size and unspecific band on the gel. So we decided to do pcr once more.</br></br></span><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of 0109, 1191, FDH containing cells.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 5ml pre-culture on 10ml tube for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer.</br></br></span><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">PCR amplification of piBR181 vector : 10 sample</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/65/120912.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">The full PCR construct of piBR181 vector is 5297bp. From the gel image, we could find that right size of piBR181 PCR product clearly exists. </br></br></span><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc & pBAD/0109 pBAD/1191 pBAD MG1655 expression check (sampling and induction)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cells (500λ of cells put into 50mL LB with 50λ Ampicillin) and checked Optical Density. According to the O.D. value, we induced 37℃ and 30℃ sample and did sampling. Each samples were induced by different conditions ; 1mM and 2mM of IPTG, 10mM and 30mM of arabinose.</br></br></span><br />
<br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Moth_0109 pBAD expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/da/120913_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we ran 10% SDS-PAGE gel and checked the presence of 59.94kDa protein, formate--tetrahydrofolate ligase. We couldn’t find any band for this protein. </br><br />
There was no significant band on the gel, so we couldn’t conclude that this protein is expressed. Our opinion is that we should run Western blot to exactly find this protein. </br></br></span><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Moth_1191 pBAD expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/af/120913-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run 15% SDS-PAGE gel and checked the presence of 33.92kDa protein, methylenetetrahydrofolate reductase. Although we were not able to discriminate each band of insoluble fraction, We could find that there exists right band at the right size. <br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pBAD expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/9e/120913-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run 8% SDS-PAGE gel and checked the presence of 40.37 kDa protein, Formate dehydrogenase. We couldn’t find any band for this protein. </br><br />
There was no significant band on the gel, so we couldn’t conclude that this protein is expressed. Our opinion is that we should run Western blot to exactly find this protein. <br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/1/1b/120913-4.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run 8% SDS-PAGE gel and checked the presence of 40.37 kDa protein, Formate dehydrogenase. We couldn’t find any band for this protein. </br><br />
There was no significant band on the gel, so we couldn’t conclude that this protein is expressed. Our opinion is that we should run Western blot to exactly find this protein. <br />
</br></br></span><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Gibson assembly – assembly of pcr fragments (pcr fragment of vector added)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Among three methods to assemble DNA fragments, we chose third method, which is assembly of PCR amplified DNA fragments and vector.<br />
0.17umoles of each fragments are added into the reaction sample, and volume to be added for each fragments has calculated on the assumption that average MW of each base is 660Da. Table below is the result of calculation.</br></br></span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/b7/120914_f1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8b/120914_f2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>After 1 hour of incubation on 50℃, the vector was transformed into TOP10 and NEB competent cell. Cells were plated and grown overnight during colony formation. </br></br></span><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Gibson assembly colony pcr</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fb/120916.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">As we couldn’t get full fragment by colony-pcr, we did 3ml culture, plasmid mini-prep and PCR for each colony. This will take one more day to get result.</br></br></span><br />
</div><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Gibson assembly fragment PCR after prep</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f2/120917.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">There appeared correct size of band with several unspecific bands on the gel. We couldn’t even try to do gel extraction because the band was too thin and faint. We decided to do more rounds of pcr amplification.</br></br></span><br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Gibson F1 PCR after prep</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/e/eb/120919-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">We did PCR amplification after prep of F1 vector, but there appeared no band on the gel. </br></br></span><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">Gibson F2 5-1~5, F1-3-1~5, F1-4-1~5</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/3/3c/120919-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">There appeared several unspecific bands on the gel and </br></br></span><br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Pre-culture of Gibson F1, F2 colonies</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">We checked that there were several colonies on the plate. 10 colonies for each fragment were picked, and culture into 3mL LB media containing 30ul Kanamycin.</br></br></span><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Vector preparation and PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">For each cultured cells, vectors were mini-prepped, and pcr was done with proper primers (F1 : 1201 F, 1197R, F2 : 1198F, 1516R)</br></br></span><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
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Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T03:55:27Z
<p>Mandie: </p>
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<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/5e/120903.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no band for 1516 gene. Band of all three genes appeared at the right size.</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></br><br />
We, </br><br />
1. changed the fraction of primers down to 0.5ul each, (lane 1)</br><br />
2. changed the polymerase to pfu-x, (lane3)</br><br />
3. used hybrid primers ( TOPO forward, Gibson reverse : lane 4 and TOPO reverse, Gibson forward : lane 5)</br><br />
Lane 2 was the control experiment with Hotstartaq polymerase.</br><br />
</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of piBR181 vector containing cells. </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 3ml pre-culture on five 10ml tubes for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer. </br></br></span><br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/ba/120905_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragment, we did PCR amplification of each gene once more. Correct size of bands appeared. </br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1197</i> <i>1516</i> PCR amplification </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/94/120905-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1197 once more. And we did PCR amplification of 1516 gene once more with the same primers. Correct size of bands appeared for 1197 gene, but there was no band for 1516 gene. . </br></br></span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1198</i> <i>1516</i>PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8c/120906.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1198 once more. And we did PCR amplification of 1516 gene once more with the same primers. For 1516 gene, we couldn’t have right size of band for new primers. So we decided to use the fragments of 1516 that does not contain terminator sequence. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">Pre-culture of <i>0109</i>, <i>1191</i>, FDH containing cells.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">We did 5ml pre-culture on 10ml tube for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer.<br />
<br />
</br></br></span><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">FDH pTrc & pBAD/<i>0109</i> pBAD/<i>1191</i> pBAD MG1655 expression check (sampling and induction)</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Procedure</b></br></br><br />
<span id="little">We seeded cells (500λ of cells put into 50mL LB with 50λ Ampicillin) and checked Optical Density. According to the O.D. value, we induced 37℃ and 30℃ sample and did sampling. Each samples were induced by different conditions ; 1mM and 2mM of IPTG, 10mM and 30mM of arabinose.</br></br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">Moth_0109 expression optimization _ gel picture</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/64/120909.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 59.94kDa protein, formate--tetrahydrofolate ligase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of 0109 gene on pBAD vector has failed and decided to try once more.</br></br></span><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Moth_1191 expression optimization _ gel picture </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/e/e0/120909_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 33.92kDa protein, methylenetetrahydrofolate reductase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of 1191 gene on pBAD vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc expression optimization _ gel picture </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/bf/120909-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 40.37kDa protein, Formate dehydrogenase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of FDH gene on pTrc vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pBAD expression optimization _ gel picture</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/6d/120909-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 40.37kDa protein, Formate dehydrogenase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of FDH gene on pTrc vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Primer design to PCR-amplify piBR181 vector.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d5/1209094.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For the Gibson assembly with RE digestion of vector, we had several problems with de-phosphorylation and purification after de-phosphorylation. Therefore we decided to PCR-amplify the piBR181 vector and use the PCR product in Gibson assembly. </br></br></span><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">PCR amplification of 1202 gene with new primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/ad/120910.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">The correct size of band appeared for the second PCR product, but there was no band appeared for the first PCR product. </br></br></span><br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">PCR amplification of piBR181 vector</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/66/120911.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">The correct size of piBR181 vector pcr product is 5297bp. There appeared the right size and unspecific band on the gel. So we decided to do pcr once more.</br></br></span><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of 0109, 1191, FDH containing cells.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 5ml pre-culture on 10ml tube for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer.</br></br></span><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">PCR amplification of piBR181 vector : 10 sample</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/65/120912.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">The full PCR construct of piBR181 vector is 5297bp. From the gel image, we could find that right size of piBR181 PCR product clearly exists. </br></br></span><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc & pBAD/0109 pBAD/1191 pBAD MG1655 expression check (sampling and induction)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cells (500λ of cells put into 50mL LB with 50λ Ampicillin) and checked Optical Density. According to the O.D. value, we induced 37℃ and 30℃ sample and did sampling. Each samples were induced by different conditions ; 1mM and 2mM of IPTG, 10mM and 30mM of arabinose.</br></br></span><br />
<br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Moth_0109 pBAD expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/da/120913_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we ran 10% SDS-PAGE gel and checked the presence of 59.94kDa protein, formate--tetrahydrofolate ligase. We couldn’t find any band for this protein. </br><br />
There was no significant band on the gel, so we couldn’t conclude that this protein is expressed. Our opinion is that we should run Western blot to exactly find this protein. </br></br></span><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Moth_1191 pBAD expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/af/120913-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run 15% SDS-PAGE gel and checked the presence of 33.92kDa protein, methylenetetrahydrofolate reductase. Although we were not able to discriminate each band of insoluble fraction, We could find that there exists right band at the right size. <br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pBAD expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/9e/120913-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run 8% SDS-PAGE gel and checked the presence of 40.37 kDa protein, Formate dehydrogenase. We couldn’t find any band for this protein. </br><br />
There was no significant band on the gel, so we couldn’t conclude that this protein is expressed. Our opinion is that we should run Western blot to exactly find this protein. <br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/1/1b/120913-4.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run 8% SDS-PAGE gel and checked the presence of 40.37 kDa protein, Formate dehydrogenase. We couldn’t find any band for this protein. </br><br />
There was no significant band on the gel, so we couldn’t conclude that this protein is expressed. Our opinion is that we should run Western blot to exactly find this protein. <br />
</br></br></span><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Gibson assembly – assembly of pcr fragments (pcr fragment of vector added)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Among three methods to assemble DNA fragments, we chose third method, which is assembly of PCR amplified DNA fragments and vector.<br />
0.17umoles of each fragments are added into the reaction sample, and volume to be added for each fragments has calculated on the assumption that average MW of each base is 660Da. Table below is the result of calculation.</br></br></span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/b7/120914_f1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8b/120914_f2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>After 1 hour of incubation on 50℃, the vector was transformed into TOP10 and NEB competent cell. Cells were plated and grown overnight during colony formation. </br></br></span><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Gibson assembly colony pcr</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fb/120916.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">As we couldn’t get full fragment by colony-pcr, we did 3ml culture, plasmid mini-prep and PCR for each colony. This will take one more day to get result.</br></br></span><br />
</div><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Gibson assembly fragment PCR after prep</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f2/120917.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">There appeared correct size of band with several unspecific bands on the gel. We couldn’t even try to do gel extraction because the band was too thin and faint. We decided to do more rounds of pcr amplification.</br></br></span><br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br> <br />
<div class="note-title">Gibson F1 PCR after prep</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/e/eb/120919-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">We did PCR amplification after prep of F1 vector, but there appeared no band on the gel. </br></br></span><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">Gibson F2 5-1~5, F1-3-1~5, F1-4-1~5</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/3/3c/120919-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">There appeared several unspecific bands on the gel and </br></br></span><br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
</br></br><br />
</div><br />
</div><br />
<br />
</body><br />
</html><br />
{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:120919-2.PNG
File:120919-2.PNG
2012-10-27T03:51:32Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120919-1.PNG
File:120919-1.PNG
2012-10-27T03:50:50Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120917.PNG
File:120917.PNG
2012-10-27T03:48:45Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120916.PNG
File:120916.PNG
2012-10-27T03:48:36Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120914_f2.PNG
File:120914 f2.PNG
2012-10-27T03:43:37Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120914_f1.PNG
File:120914 f1.PNG
2012-10-27T03:42:46Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120913-4.PNG
File:120913-4.PNG
2012-10-27T03:38:55Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120913-3.PNG
File:120913-3.PNG
2012-10-27T03:38:04Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120913-2.PNG
File:120913-2.PNG
2012-10-27T03:37:58Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120913_-_1.PNG
File:120913 - 1.PNG
2012-10-27T03:37:52Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120912.PNG
File:120912.PNG
2012-10-27T03:29:57Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120911.PNG
File:120911.PNG
2012-10-27T03:26:17Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120910.PNG
File:120910.PNG
2012-10-27T03:24:39Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T03:22:23Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
{{:Team:KAIST_Korea/navi}}<br />
<html><br />
<head><br />
<link href='http://fonts.googleapis.com/css?family=Gloria+Hallelujah|Open+Sans:600italic,400,800' rel='stylesheet' type='text/css'><br />
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text-align:justify;<br />
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<br />
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}<br />
#kaistcontent ol<br />
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list-style-type:decimal;<br />
padding: 0px 0px 0px 30px;<br />
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<br />
#kaistcontent #sub-sub-title<br />
{<br />
font-family: 'Open Sans', sans-serif;<br />
color:#848383;<br />
font-weight:bold;<br />
font-size:14pt;<br />
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background-image:url('https://static.igem.org/mediawiki/2012/a/a0/KAIST_labnote_logo.png');<br />
background-repeat:no-repeat;<br />
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</style><br />
</head><br />
<br />
<body><br />
<div id="main"><br />
<img id="top-img" src="https://static.igem.org/mediawiki/2012/6/6d/Back_home.PNG"/><br />
<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
</div><br />
<br />
<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/5e/120903.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no band for 1516 gene. Band of all three genes appeared at the right size.</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></br><br />
We, </br><br />
1. changed the fraction of primers down to 0.5ul each, (lane 1)</br><br />
2. changed the polymerase to pfu-x, (lane3)</br><br />
3. used hybrid primers ( TOPO forward, Gibson reverse : lane 4 and TOPO reverse, Gibson forward : lane 5)</br><br />
Lane 2 was the control experiment with Hotstartaq polymerase.</br><br />
</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of piBR181 vector containing cells. </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 3ml pre-culture on five 10ml tubes for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer. </br></br></span><br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/ba/120905_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragment, we did PCR amplification of each gene once more. Correct size of bands appeared. </br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1197</i> <i>1516</i> PCR amplification </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/94/120905-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1197 once more. And we did PCR amplification of 1516 gene once more with the same primers. Correct size of bands appeared for 1197 gene, but there was no band for 1516 gene. . </br></br></span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1198</i> <i>1516</i>PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8c/120906.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1198 once more. And we did PCR amplification of 1516 gene once more with the same primers. For 1516 gene, we couldn’t have right size of band for new primers. So we decided to use the fragments of 1516 that does not contain terminator sequence. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">Pre-culture of <i>0109</i>, <i>1191</i>, FDH containing cells.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">We did 5ml pre-culture on 10ml tube for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer.<br />
<br />
</br></br></span><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">FDH pTrc & pBAD/<i>0109</i> pBAD/<i>1191</i> pBAD MG1655 expression check (sampling and induction)</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Procedure</b></br></br><br />
<span id="little">We seeded cells (500λ of cells put into 50mL LB with 50λ Ampicillin) and checked Optical Density. According to the O.D. value, we induced 37℃ and 30℃ sample and did sampling. Each samples were induced by different conditions ; 1mM and 2mM of IPTG, 10mM and 30mM of arabinose.</br></br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">Moth_0109 expression optimization _ gel picture</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/64/120909.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 59.94kDa protein, formate--tetrahydrofolate ligase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of 0109 gene on pBAD vector has failed and decided to try once more.</br></br></span><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Moth_1191 expression optimization _ gel picture </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/e/e0/120909_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 33.92kDa protein, methylenetetrahydrofolate reductase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of 1191 gene on pBAD vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc expression optimization _ gel picture </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/bf/120909-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 40.37kDa protein, Formate dehydrogenase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of FDH gene on pTrc vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pBAD expression optimization _ gel picture</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/6d/120909-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 40.37kDa protein, Formate dehydrogenase. The gel was ruined during staining/destaining procedure, and we couldn’t find any band for this protein. </br><br />
Therefore, we concluded that expression of FDH gene on pTrc vector has failed and decided to try once more.<br />
</br></br></span><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pBAD expression optimization _ gel picture</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d5/1209094.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">For the Gibson assembly with RE digestion of vector, we had several problems with de-phosphorylation and purification after de-phosphorylation. Therefore we decided to PCR-amplify the piBR181 vector and use the PCR product in Gibson assembly. </br></br></span><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
</br></br><br />
</div><br />
</div><br />
<br />
</body><br />
</html><br />
{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:1209094.PNG
File:1209094.PNG
2012-10-27T03:20:47Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120909-3.PNG
File:120909-3.PNG
2012-10-27T03:19:47Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120909-2.PNG
File:120909-2.PNG
2012-10-27T03:19:38Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120909_-_1.PNG
File:120909 - 1.PNG
2012-10-27T03:19:25Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120909.PNG
File:120909.PNG
2012-10-27T03:12:04Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T03:06:53Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
{{:Team:KAIST_Korea/navi}}<br />
<html><br />
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margin-left:auto;<br />
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list-style-type:decimal;<br />
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font-family: 'Open Sans', sans-serif;<br />
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font-size:14pt;<br />
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background-image:url('https://static.igem.org/mediawiki/2012/a/a0/KAIST_labnote_logo.png');<br />
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<br />
<body><br />
<div id="main"><br />
<img id="top-img" src="https://static.igem.org/mediawiki/2012/6/6d/Back_home.PNG"/><br />
<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
</div><br />
<br />
<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/5e/120903.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no band for 1516 gene. Band of all three genes appeared at the right size.</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></br><br />
We, </br><br />
1. changed the fraction of primers down to 0.5ul each, (lane 1)</br><br />
2. changed the polymerase to pfu-x, (lane3)</br><br />
3. used hybrid primers ( TOPO forward, Gibson reverse : lane 4 and TOPO reverse, Gibson forward : lane 5)</br><br />
Lane 2 was the control experiment with Hotstartaq polymerase.</br><br />
</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of piBR181 vector containing cells. </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 3ml pre-culture on five 10ml tubes for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer. </br></br></span><br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/ba/120905_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragment, we did PCR amplification of each gene once more. Correct size of bands appeared. </br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1197</i> <i>1516</i> PCR amplification </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/94/120905-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1197 once more. And we did PCR amplification of 1516 gene once more with the same primers. Correct size of bands appeared for 1197 gene, but there was no band for 1516 gene. . </br></br></span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1198</i> <i>1516</i>PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8c/120906.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1198 once more. And we did PCR amplification of 1516 gene once more with the same primers. For 1516 gene, we couldn’t have right size of band for new primers. So we decided to use the fragments of 1516 that does not contain terminator sequence. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">Pre-culture of <i>0109</i>, <i>1191</i>, FDH containing cells.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Discussion</b></br></br><br />
<span id="little">We did 5ml pre-culture on 10ml tube for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer.<br />
<br />
</br></br></span><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title">FDH pTrc & pBAD/<i>0109</i> pBAD/<i>1191</i> pBAD MG1655 expression check (sampling and induction)</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Procedure</b></br></br><br />
<span id="little">We seeded cells (500λ of cells put into 50mL LB with 50λ Ampicillin) and checked Optical Density. According to the O.D. value, we induced 37℃ and 30℃ sample and did sampling. Each samples were induced by different conditions ; 1mM and 2mM of IPTG, 10mM and 30mM of arabinose.<br />
<br />
</br></br></span><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
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Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T03:03:18Z
<p>Mandie: </p>
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<img id="top-img" src="https://static.igem.org/mediawiki/2012/6/6d/Back_home.PNG"/><br />
<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
</div><br />
<br />
<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/5e/120903.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no band for 1516 gene. Band of all three genes appeared at the right size.</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></br><br />
We, </br><br />
1. changed the fraction of primers down to 0.5ul each, (lane 1)</br><br />
2. changed the polymerase to pfu-x, (lane3)</br><br />
3. used hybrid primers ( TOPO forward, Gibson reverse : lane 4 and TOPO reverse, Gibson forward : lane 5)</br><br />
Lane 2 was the control experiment with Hotstartaq polymerase.</br><br />
</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of piBR181 vector containing cells. </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 3ml pre-culture on five 10ml tubes for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer. </br></br></span><br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/ba/120905_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragment, we did PCR amplification of each gene once more. Correct size of bands appeared. </br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1197</i> <i>1516</i> PCR amplification </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/94/120905-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1197 once more. And we did PCR amplification of 1516 gene once more with the same primers. Correct size of bands appeared for 1197 gene, but there was no band for 1516 gene. . </br></br></span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1198</i> <i>1516</i>PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8c/120906.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1198 once more. And we did PCR amplification of 1516 gene once more with the same primers. For 1516 gene, we couldn’t have right size of band for new primers. So we decided to use the fragments of 1516 that does not contain terminator sequence. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
</br></br><br />
</div><br />
</div><br />
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</body><br />
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{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:120906.PNG
File:120906.PNG
2012-10-27T03:02:45Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T03:01:11Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
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<br />
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<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
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<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/5e/120903.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no band for 1516 gene. Band of all three genes appeared at the right size.</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></br><br />
We, </br><br />
1. changed the fraction of primers down to 0.5ul each, (lane 1)</br><br />
2. changed the polymerase to pfu-x, (lane3)</br><br />
3. used hybrid primers ( TOPO forward, Gibson reverse : lane 4 and TOPO reverse, Gibson forward : lane 5)</br><br />
Lane 2 was the control experiment with Hotstartaq polymerase.</br><br />
</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of piBR181 vector containing cells. </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 3ml pre-culture on five 10ml tubes for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer. </br></br></span><br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></span><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/ba/120905_-_1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To amplify the amount of each fragment, we did PCR amplification of each gene once more. Correct size of bands appeared. </br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1197</i> <i>1516</i> PCR amplification </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/94/120905-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">To amplify the amount of each fragments, we did PCR amplification of 1197 once more. And we did PCR amplification of 1516 gene once more with the same primers. Correct size of bands appeared for 1197 gene, but there was no band for 1516 gene. . </br></br></span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
</br></br><br />
</div><br />
</div><br />
<br />
</body><br />
</html><br />
{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:120905-2.PNG
File:120905-2.PNG
2012-10-27T02:57:56Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120905_-_1.PNG
File:120905 - 1.PNG
2012-10-27T02:57:23Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T02:49:39Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
{{:Team:KAIST_Korea/navi}}<br />
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background-image:url('https://static.igem.org/mediawiki/2012/a/a0/KAIST_labnote_logo.png');<br />
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</head><br />
<br />
<body><br />
<div id="main"><br />
<img id="top-img" src="https://static.igem.org/mediawiki/2012/6/6d/Back_home.PNG"/><br />
<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
</div><br />
<br />
<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/5e/120903.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no band for 1516 gene. Band of all three genes appeared at the right size.</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/c/c1/120903-%2B--.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">Also for this time, there was no right size of band appeared for 1516 gene. </br></br><br />
We, </br><br />
1. changed the fraction of primers down to 0.5ul each, (lane 1)</br><br />
2. changed the polymerase to pfu-x, (lane3)</br><br />
3. used hybrid primers ( TOPO forward, Gibson reverse : lane 4 and TOPO reverse, Gibson forward : lane 5)</br><br />
Lane 2 was the control experiment with Hotstartaq polymerase.</br><br />
</br></span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of piBR181 vector containing cells. </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We did 3ml pre-culture on five 10ml tubes for the cells containing each gene. Cell was grown about 16hours after pre-culture, and then induced with proper inducer. </br></br></span><br />
<br />
</div><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
</br></br><br />
</div><br />
</div><br />
<br />
</body><br />
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{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:120903-%2B--.PNG
File:120903-+--.PNG
2012-10-27T02:44:53Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:%EC%BA%A1%EC%B2%98.PNG
File:캡처.PNG
2012-10-27T02:43:04Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T02:30:48Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
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<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
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<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
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<br />
<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> <i>1201</i> <i>1197</i> <i>1198</i> <i>1516</i> PCR for Gibson assembly.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/5e/120903.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no band for 1516 gene. Band of all three genes appeared at the right size.</br></br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
</br></br><br />
</div><br />
</div><br />
<br />
</body><br />
</html><br />
{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:120903.PNG
File:120903.PNG
2012-10-27T02:29:14Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_9
Team:KAIST Korea/Notebook Labnote/2012 9
2012-10-27T02:26:41Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
{{:Team:KAIST_Korea/navi}}<br />
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border-color:#EEE;<br />
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margin:0px 0px 35px 0px;<br />
background-image:url('https://static.igem.org/mediawiki/2012/a/a0/KAIST_labnote_logo.png');<br />
background-repeat:no-repeat;<br />
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<br />
<body><br />
<div id="main"><br />
<img id="top-img" src="https://static.igem.org/mediawiki/2012/6/6d/Back_home.PNG"/><br />
<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
</div><br />
<br />
<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-September</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">September</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">September 1<sup>st</sup> 2012</div></br><br />
<br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> fdh pBAD, fdh pTrc colony PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/5/55/120901.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">To check the presence of FDH gene, we picked the colony from the plate and did colony PCR. For all the colonies we picked, we confirmed that correct size of FDH is present. </br></span><br />
<br />
</div><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD cloning with remaining DNAs</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Remaining Bxb1_GTG insert was cloned into pBADmycHisC vector. And electrotransformed into MG1655.</span><br />
</br></br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="2"><br />
<div class="date">September 2<sup>nd</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc miniprep for double transformation, Bxb1_GTG_pBAD colony inoculation and vector miniprep NcoI single cut check.</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0902Fig1" src="https://static.igem.org/mediawiki/2012/1/1c/KAIST_2_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">All the vectors showed right sizes. But Bxb1_GTG_pBAD colony #1 through 3 have undesired bands. They may be the supercoiled vector.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_ATG_pTrc, pBAD, Bxb1_GTG_pTrc double transformation into pPoC and pPoCpi</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="3"><br />
<div class="date">September 3<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 double transformants vector miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0903Fig1" src="https://static.igem.org/mediawiki/2012/1/18/KAIST_3_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">Seemed to every colonies have appropriate vectors.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">September 4<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">September 5<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">September 6<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">September 7<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="8"><br />
<div class="date">September 8<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">September 9<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">September 10<sup>th</sup> 2012</div></br><br />
<br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD single cut check with different enzyme</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0910Fig1" src="https://static.igem.org/mediawiki/2012/9/98/KAIST_10_Sept.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">No desired size(5.6kb) vector appeared.</span><br />
<br />
</br<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">September 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">September 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1_GTG_pBAD colony #4~7 inoculation, miniprep and single cut check<br />
Single cut with SnaBI<br />
</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Single cut with SnaBI</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0912Fig1" src="https://static.igem.org/mediawiki/2012/1/13/KAIST_12_Sept.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Cloning completed</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">September 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">September 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">September 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062 transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Templates for Auto-regulated FlipFlop project.</span><br />
</br></br><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">September 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_C0060, BBa_C0061, BBa_C0062, bFMO inoculation and enzyme cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_C0060, BBa_C0061, BBa_C0062: from colony, EcoRI single cut<br><br><br />
<ul style="list-style-type:square;padding:0px 0px 0px 40px"><br />
<li> BBa_C0060: 2859bp </li><br />
<li> BBa_C0061: 2688bp </li><br />
<li> BBa_C0062: 2826bp </li><br />
<li>bFMO: from cell stock, EcoRI single cut</li><br />
</ul></span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0916#Fig#1" src="https://static.igem.org/mediawiki/2012/0/0f/16_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">September 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
</br><br />
<span id="little">Promoter construct: 210bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0917Fig1" src="https://static.igem.org/mediawiki/2012/d/d3/KAIST_17_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">Promoter PCR failed</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0917Fig2" src="https://static.igem.org/mediawiki/2012/d/d7/KAIST_17_Sept_2.JPG"></img><br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">September 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0918Fig1" src="https://static.igem.org/mediawiki/2012/5/54/KAIST_18_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Terminator PCR failed</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">September 19<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Terminator: 179bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0919Fig1" src="https://static.igem.org/mediawiki/2012/4/44/KAIST_19_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">September 20<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation for OE PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 Xis: 777bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0920Fig1" src="https://static.igem.org/mediawiki/2012/8/8e/KAIST_20_Sept.JPG"></img></div><br />
<div style="clear:both;"></div></br><br />
<span id="little">bFMO PCR w/ various condtions</span></br><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0920Fig2" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_20_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1_Xis-pGEM_B1 transfomation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene – Bxb1_Xis arrived and transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">September 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">September 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Overlapping extension PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0922Fig1" src="https://static.igem.org/mediawiki/2012/f/f5/KAIST_22_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"> → Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">September 23<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR-Nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig1" src="https://static.igem.org/mediawiki/2012/4/41/KAIST_23_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<b>Discussions</b></br></br><br />
<span id="little">There was too little full construct nested. We think this is due to primer interfering by its LVA homology.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0923Fig2" src="https://static.igem.org/mediawiki/2012/a/ae/KAIST_23_Sept_2.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">Bands were more intense than those of first trial.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">September 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0924Fig1" src="https://static.igem.org/mediawiki/2012/6/6e/KAIST_24_Sept.JPG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">P1, PI construct </div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Full fragment for pAuto is overlapped. The right size(3794bp) fragments are gel extracted. </br></br><br />
<br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/4/41/120924-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
</div><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">September 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">September 26<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Full pAuto and pAutoSimple Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/21/120926-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">The band intensity is too weak.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/04/120926-2-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Full construct(2.5kb) for pAutoIntegrase is overlapped. The products are pcr purified and performed nested-pcr. 4th lane is the OE pcr product with rearranged template concentration. </br></br><br />
</span><br />
<br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/06/120926-2-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Template preparation of terminator</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fd/120926-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Terminator template for OE pcr is re-amplified with pfu-X DNA polymerase. The products are pcr purified. </br><br />
</span><br />
<br />
</div><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">September 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">September 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">September 29<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template preparation of bFMO (for pAuto and pAutoSimple)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/f8/120929-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>To find out the right temperature for primer binding, we performed gradient pcr from 45℃~55℃.<br />
But, all the pcr products showed non-specific binding of primers, and we selected the most proper temperature.<br />
</br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Overlapping extension PCR Gel Extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7d/120930.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
</br></br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">September 30<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Template vector preparation – double digestion check with EcoRI and PstI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/0/0a/120930-..PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Vector(BBa_J04450-pSB4A5) which will be used for cloning of pAutoIntegrase.</br></span><br />
<br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">bFMO template amplification with optimized condition</div><br />
<br />
<b>Results</b></br></br><br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">pAutoIntegrase cloning check</div><br />
<b>Results</b></br></br><br />
<span id="little">PCR amplified from prepped vector </br><br />
</span><br />
<br />
<div id="content_note" ><br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
</br></br><br />
</div><br />
</div><br />
<br />
</body><br />
</html><br />
{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:120901.PNG
File:120901.PNG
2012-10-27T02:25:35Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_8
Team:KAIST Korea/Notebook Labnote/2012 8
2012-10-27T02:20:56Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
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<br />
<body><br />
<div id="main"><br />
<img id="top-img" src="https://static.igem.org/mediawiki/2012/6/6d/Back_home.PNG"/><br />
<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
</div><br />
<br />
<div id="top-img-description-box"><span id="top-img-description">Notebook : Labnote-August</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">August</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">August 1<sup>st</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">PCR amplification of <i>Moth_1191</i>, <i>1199</i> gene : decreasing annealing temperature down to 45℃</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">To make sure there’s no problem with unspecific binding or annealing temperature, we decreased annealing condition down to 45℃.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0801Fig1" src="https://static.igem.org/mediawiki/2012/e/e1/KAIST_08012012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">There was no band and it became certain that there are no problems with annealing procedure.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Induction of <i>Moth_0109</i> gene (sampling) and running SDS-PAGE gel. </div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">We induced <i>Moth_0109</i> gene in pBAD/myc vector with different Arabinose conditions and temperature: 10mM, 30mM and 37℃, 30℃. And then, we run the sample into SDS-PAGE gel. The gel is stained with coomassie blue for 1 hour and de-stained for overnight.</span><br />
</br></br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">PCR amplification of <i>Moth_1191</i>, <i>1199</i> gene with Gibson and TOPO primer</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">To see whether there are any problem with Gibson primers that we’ve designed, we did one more pcr amplification. Primers for Gibson assembly and TOPO cloning are mixed, and we could see any problems with forward or reverse primers. </span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0801Fig2" src="https://static.igem.org/mediawiki/2012/6/62/KAIST_08012012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Each lane indicates:</br></br><br />
<ul style="list-style-type:square"><br />
<li><i>Moth_1191_1</i> : TOPO forward primer and Gibson reverse primer</li> <br />
<li><i>Moth_1191_2</i> : Gibson forward primer and TOPO reverse primer</li><br />
<li><i>Moth_1199_1</i> : Gibson forward primer and TOPO reverse primer</li><br />
<li><i>Moth_1199_2</i> : TOPO forward primer and Gibson reverse primer</li><br />
</ul><br />
</br><br />
Correct size of band appeared when topo forward primer and Gibson reverse primer was used in amplification of both genes. Therefore, we can conclude that there is some unknown problem with forward primers.</span><br />
<br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi OE PCR with changed primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0801Fig1" src="https://static.igem.org/mediawiki/2012/8/81/KAIST_2nd_08012012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">→ Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="2"><br />
<div class="date">August 2<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Primer design for FDH derived from <i>Candida boidinii</i> mutant</div><br />
<br />
<div id="content_note" ><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0802Fig1" src="https://static.igem.org/mediawiki/2012/b/b8/KAIST_08022012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">These primers are universial primer that can be used for insertion of this gene into both pTrcHis2A vector and pBAD/mycHisC vector. </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Expression check of <i>Moth_0109</i> gene on pBAD/mycHisC vector (Result)</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 59.94 kDa protein, formate tetrahydrofolate ligase. </span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0802Fig2" src="https://static.igem.org/mediawiki/2012/7/7c/KAIST_08022012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">The band of PAGE gel was not certain, so we </span><br />
<!--이것도 쓰다가 말았음;; --><br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi cloning - Digestion</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Restriction enzyme digestion</span><br />
</br></br><br />
<span><br />
<ul style="padding:0px 0px 0px 40px;"><br />
<li>Enzyme pre-mix / each reaction – 37℃ 1hr, 80℃ 20min</li></br><br />
<div align="center"><img id="figure" alt="2_0802Fig1" src="https://static.igem.org/mediawiki/2012/1/1e/KAIST_2nd_08022012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<li>Digestion mixture – 37℃ 1hr, 80℃ 20min</li></br><br />
<div align="center"><img id="figure" alt="2_0802Fig2" src="https://static.igem.org/mediawiki/2012/7/79/KAIST_2nd_08022012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</ul><br />
</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1 integrase transformation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene arrived.</br></span><br />
<span id="little">Transformed into DH5α </span><br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<section id="3"><br />
<div class="date">August 3<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 integrase-DH5α streaking</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">Synthesized gene was cloned into pGEM-B1 vector. It seems the vector concentration is too high so that too much colonies may appeared.</span><br />
<br />
</br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi cloning - Ligation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Ligation - 16℃ overnight</span><br />
</br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0803Fig3" src="https://static.igem.org/mediawiki/2012/8/81/KAIST_2nd_08032012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Transformed into DH5α chemically competent cell.</span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">BBa_J04450 transformation into DH5α</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">For pSB1C3 vector amplification</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<section id="4"><br />
<div class="date">August 4<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title"><i>1191</i> and <i>1199</i> forward primer design</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">We designed new forward primers for both <i>1191</i> and <i>1199</i> gene that are thought to solve problems appeared during PCR amplification.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0804Fig1" src="https://static.igem.org/mediawiki/2012/5/59/KAIST_08042012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_J04450 and Bxb1-pGEM colony picking and inoculation.</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Made master plates and inoculated in 3mL of LB.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="5"><br />
<div class="date">August 5<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="6"><br />
<div class="date">August 6<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_J04450 Plasmid miniprep</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_J04450(pSB1C3): 285.8ng/uL</span></br><br />
<span id="little">purity:2.10</span><br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Requested part arrived</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Each parts were streaked on appropriate plates.</br></br><br />
<ul><br />
<li>BBa_I11020</li><br />
<li>BBa_I11022</li><br />
<li>BBa_I11023</li><br />
<li>BBa_I11030</li><br />
<li>BBa_K137007</li><br />
<li>BBa_K137008</li><br />
<li>BBa_K137010</li><br />
<li>BBa_K112001</li><br />
<li>BBa_K112141</li><br />
<li>BBa_K112142</li><br />
</ul><br />
</span><br />
</br></br><br />
<br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">BBa_K381001(GFP) and BBa_K398005(pSB1C3) transformation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_K381001(GFP): for GFP control</span></br><br />
<span id='little'>BBa_K398005(pSB1C3): for pSB1C3 template</span></br><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="7"><br />
<div class="date">August 7<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">FDH gene PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0807Fig1" src="https://static.igem.org/mediawiki/2012/7/7b/KAIST_08072012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">We amplified 2 samples of formate dehydrogenase gene with the primers we designed. From gel electrophoresis picture, we can say that oligonucleotides with the proper size 1100bp are present in the sample.</span><br />
<div align="center"><img id="figure" alt="0807Fig2" src="https://static.igem.org/mediawiki/2012/e/eb/KAIST_08072012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br> <br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_J04450(pSB1C3) enzyme cut check(single cut with EcoRI)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0807Fig1" src="https://static.igem.org/mediawiki/2012/9/93/KAIST_2nd_08072012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Vector was double digested. The insert and vector sizes were correct</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi OE PCR product nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0807Fig2" src="https://static.igem.org/mediawiki/2012/9/91/KAIST_2nd_08072012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">→ Gel extracted</span><br />
<br />
</br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">BBa_K398005 Plasmid miniprep & single cut check with EcoRI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0807Fig3" src="https://static.igem.org/mediawiki/2012/c/c9/KAIST_2nd_08072012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pPoC insert</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0807Fig4" src="https://static.igem.org/mediawiki/2012/d/d5/KAIST_2nd_08072012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"><br />
<ul style="list-style-type:square; padding:0px 0px 0px 40px;"><br />
<li>HSTaq Pol MM : 25uL</li><br />
<li>DW : 1uL</li><br />
<li>templates : 22uL</li><br />
<li>primers : 2uL</li><br />
<br />
</ul><br />
</span></br></br><br />
<br />
<span id="little">pPoCpi insert</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0807Fig4" src="https://static.igem.org/mediawiki/2012/d/d5/KAIST_2nd_08072012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"><br />
<ul style="list-style-type:square; padding:0px 0px 0px 40px;"><br />
<li>HSTaq Pol MM : 25uL</li><br />
<li>templates : 23.2uL</li><br />
<li>primers : 2uL</li><br />
</ul><br />
</span><br />
</br><br />
</br><br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="2_0807Fig5" src="https://static.igem.org/mediawiki/2012/e/ea/KAIST_2nd_08072012_Fig5.png"></img></div><br />
<div style="clear:both;"></div><br />
</br><br />
<span id ="little">→ Concentration after gel extraction was not sufficient for cloning</span></br></br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping PCR template preparation</div><br />
<br />
<div id="content_note" ><br />
<br />
<div align="center"><img id="figure" alt="2_0807Fig6" src="https://static.igem.org/mediawiki/2012/9/99/KAIST_2nd_08072012_Fig6.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0807Fig7" src="https://static.igem.org/mediawiki/2012/3/3a/KAIST_2nd_08072012_Fig7.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<br />
<br />
<section id="8"><br />
<div class="date">August 8<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">FDH gene and pTrcHis2A, pBAD/mycHisC vector enzyme digestion</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We mixed restriction enzymes and buffers with DNA samples as shown below.</span></br></br><br />
<br />
<div align="center"><img id="figure" alt="0808Fig1" src="https://static.igem.org/mediawiki/2012/3/31/KAIST_08082012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<span id="little">After mixing the enzymes and buffer with the sample, we incubated the sample on 37℃ for 1 hour and then on 65℃ for 20minutes. </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pTrcHis2A, pBAD/myc vector dephosphorylation</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<span id="little">We mixed phosphatidase and buffer with DNA sample as shown below.</span></br></br><br />
<div align="center"><img id="figure" alt="0808Fig2" src="https://static.igem.org/mediawiki/2012/4/49/KAIST_08082012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">After mixing the enzyme and buffer with the sample, we incubated the sample on 37℃ for 1 hour 20minutes and then on 70℃ for 5minutes. </span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">ligation of FDH gene with pTrcHis2A and pBAD/mycHisC vectors</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<span id="little">We mixed ligase and buffer with DNA sample as shown below.</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="0808Fig3" src="https://static.igem.org/mediawiki/2012/4/4e/KAIST_08082012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">After mixing the enzyme and buffer with the sample, we incubated the sample on 16℃ for 16 hours.</span><br />
<br />
</br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Primer amplification of <i>1191</i> and <i>1199</i></div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0808Fig4" src="https://static.igem.org/mediawiki/2012/5/5d/KAIST_08082012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pPoC insert</span></br></br><br />
<div align="center"><img id="figure" alt="2_0808Fig2" src="https://static.igem.org/mediawiki/2012/9/9b/KAIST_2nd_08082012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">pPoCpi insert</span></br></br><br />
<div align="center"><img id="figure" alt="2_0808Fig3" src="https://static.igem.org/mediawiki/2012/5/53/KAIST_2nd_08082012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<span><br />
<ul style="list-style-type:square"><br />
<br />
<li>HSTaq Pol MM : 25uL</li><br />
<li>DW : 5.2uL</li><br />
<li>templates : 19.8uL</li><br />
<li>primers : 2uL</li><br />
<li>pfu-x : 0.1uL</li><br />
<br />
</ul><br />
</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0808Fig1" src="https://static.igem.org/mediawiki/2012/3/31/KAIST_2nd_08082012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">→ Gel extraction</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping PCR template preparation</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0808Fig4" src="https://static.igem.org/mediawiki/2012/5/5d/KAIST_2nd_08082012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<br />
<section id="9"><br />
<div class="date">August 9<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">vector transformation into TOP10 cell</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Transformation of pTrcHis2A vector into TOP10 cell was successful. So we did vector mini-preparation after the transformation. However, transformation of pBAD vector has failed, so we ligated the samples again. </span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0809Fig1" src="https://static.igem.org/mediawiki/2012/e/e9/KAIST_08092012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">This is gel picture to confirm that FDH is well inserted into TOP10 cells. 3 wells on the left are PCR product of <!--??/--></span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 integrase cloning into pTrcHis2a and pBADmycHisC</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 integrase: 1515bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0809Fig1" src="https://static.igem.org/mediawiki/2012/3/33/KAIST_2nd_08092012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">→ Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Extracted OE PCR product cloning into pSB1C3</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pSB1C3 was originated from BBa_K398005.</br></br>Transformed into TOP10</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<span id="little">Transformed colony was grown in LB media. No colony shows fluorescence. </span><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">We think cloning failed.</span><br />
<br />
</br></br><br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="10"><br />
<div class="date">August 10<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title"> Bxb1-pTrc, pBAD colony inoculation</div><br />
<br />
<div id="content_note" ><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">In this time, PCR was done first 18 cycles without any primer.</br><br />
After that, we add 1uL of primers. And then PCR was done 20 cycles more.<br />
</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0810Fig1" src="https://static.igem.org/mediawiki/2012/7/79/KAIST_2nd_08102012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">With this OE PCR conditions, yield was better than previous conditions.</span><br />
<br />
</br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pPoC, pPoCpi insert cloning into pSB1C3</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pSB1C3 was derived from BBa_K398005<br />
</span><br />
</br></br><br />
<br />
<span><br />
<ol style="padding:0px 0px 0px 40px;"><br />
<li>Digestion – 37℃ 1hr, 80℃ 20min</li><br />
<div align="center"><img id="figure" alt="2_0810Fig5" src="https://static.igem.org/mediawiki/2012/5/5d/KAIST_2nd_08102012_Fig5.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<li>Dephosphorylation – 37℃ 30min, 80℃ 10min</li><br />
<div align="center"><img id="figure" alt="2_0810Fig6" src="https://static.igem.org/mediawiki/2012/8/87/KAIST_2nd_08102012_Fig6.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<li>Ligation – 37℃ 2hr</li><br />
<div align="center"><img id="figure" alt="2_0810Fig7" src="https://static.igem.org/mediawiki/2012/f/fb/KAIST_2nd_08102012_Fig7.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</ol><br />
</span><br />
</br></br><br />
<span id="little">Ligated products were transformed into DH5α.</span><br />
<br />
<br />
</br><br />
</div><br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Transformed cell inoculation and colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0810Fig8" src="https://static.igem.org/mediawiki/2012/3/3c/KAIST_2nd_08102012_Fig8.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">only pPoCpi construct cloning was completed </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="11"><br />
<div class="date">August 11<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">pTrcHis2A vector transformation into <i>E.coli</i> MG1655 cell</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pBAD/mycHisC-FDH transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 pTrcHis2A, pBADmycHisC plasmid DNA mini-prep</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">single cut check</span></br></br><br />
<div align="center"><img id="figure" alt="2_0811Fig1" src="https://static.igem.org/mediawiki/2012/d/db/KAIST_2nd_08112012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<span id="little">nested PCR to prepare more insert DNA</span></br></br><br />
<div align="center"><img id="figure" alt="2_0811Fig2" src="https://static.igem.org/mediawiki/2012/1/11/KAIST_2nd_08112012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<br />
<section id="12"><br />
<div class="date">August 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">colony PCR check of pTrcHis2A inserted FDH gene</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0812Fig1" src="https://static.igem.org/mediawiki/2012/8/80/KAIST_08122012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">We checked the presence of FDH gene by conducting colony PCR. Correct size of band appeared at about 1115bp, which means formate dehydrogenase is inserted correctly. </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pBAD FDH MG1655 electroporation</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<br />
<span id="little">Fail</span><br />
<!--여기 쓸말 더 있으면 써줘요 지환오빠 ㅋㅋ--><br />
</br><br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="13"><br />
<div class="date">August 13<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">FDH pBAD MG1655 electroporation</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cells on 7:10 AM and did cell-down on 10:00AM. And then we plated cells on 1:00PM and checked colony on 8/14 5:00AM</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<!-- <br />
<div align="center"><img id="figure" alt="08##Fig#" src="#"></img></div><br />
<div style="clear:both;"></div> Colony pic.--><br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc MG1655 expression check (sampling and induction)</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cells (500λ of cells put into 50mL LB with 50λ Ampicillin) on 7 : 30 AM and checked Optical Density on 10:30pm. According to the O.D. value, we induced 37℃ sample on 10:30PM and did sampling on 4:30pm. Also, we induced 30℃ sample on 11:30pm and did sampling on 5:30pm. (6 hour induction) Each temperature samples were induced by different conditions ; 2mM and 3mM of IPTG.</span><br />
</br></br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoCpi cloning into pSB1C3</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Cloning was done by remaining DNA samples. </span><br />
</br></br><br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="14"><br />
<div class="date">August 14<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">colony PCR check of pBADmycHisC inserted FDH gene</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0814Fig1" src="https://static.igem.org/mediawiki/2012/e/e2/KAIST_08142012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">We checked the presence of FDH gene by conducting colony PCR. Correct size of band appeared at about 1115 bp, which means formate dehydrogenase is inserted correctly. </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Expression check of Formate dehydrogenase on pTrcHis2A vector (Result)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0814Fig2" src="https://static.igem.org/mediawiki/2012/6/64/KAIST_08142012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 43.7 kDa protein, formate dehydrogenase. The band appeared significantly on the correct size. </span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">FDH pBAD MG1655 seeding for expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cell on 7:00PM, so we’ll incubate cell about 16hours. (This means we have to do next seeding on 11:00AM. )</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Primer design for the insertion of <i>1202</i> and <i>0109</i></div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0814Fig3" src="https://static.igem.org/mediawiki/2012/b/bd/KAIST_08142012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">To eliminate uncertain spacer between genes, we inserted </span><br />
<!--쓰다가 말았네;;--><br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 pTrcHis2A and pPoCpi ligated products transformation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">The products were transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="15"><br />
<div class="date">August 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">FDH pBAD MG1655 expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cells on 11 : 30 AM and checked Optical Density on 1:30pm. According to the O.D. value, we induced 37℃ sample on 1:40pm and did sampling on 7:40pm. Also, we induced 30℃ sample on 3:30pm and did sampling on 9:30pm. (6 hour induction) Each temperature samples were induced by different conditions: 10mM of arabinose and 30mM of arabinose.</span><br />
<br />
</br><br />
<br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoCpi colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_08##Fig#" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_2nd_08152012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Among colonies #1~10, four colonies were inoculated.<br />
#1, 3, 4, 7<br />
</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1 pTrcHis2A colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0815Fig2" src="https://static.igem.org/mediawiki/2012/9/9d/KAIST_2nd_08152012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Among colonies #1~20, three colonies were inoculated.<br />
#2, 5, 19<br />
</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<section id="16"><br />
<div class="date">August 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">PCR amplification of <i>1201</i>, <i>1202</i>, <i>1203</i>, <i>1198</i> and <i>1191</i></div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0816Fig1" src="https://static.igem.org/mediawiki/2012/8/86/KAIST_08152012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">Primers arrived, and we began to do additional PCR amplification on the preparation for Gibson assembly. Each PCR sample has the correct size of oligonucleotides except <i>Moth 1202</i> gene.</br></br><br />
<ul><br />
<li><i>1191</i> : 921bp</br></li><br />
<li><i>1198</i> : 972bp + about 400bp = 1300 bp </br></li><br />
<li><i>1201</i> : 1341bp + ?</br></li><br />
<li><i>1202</i> : 2190bp + </br></li><br />
<li><i>1203</i> : 2025bp + 1600bp = 3625bp</br></li><br />
</ul><br />
</br><br />
We did PCR purification on the PCR product and the concentration/purity result was as shown below. <br />
</span><br />
</br></br><br />
<br />
<div align="center"><img id="figure" alt="0816Fig2" src="https://static.igem.org/mediawiki/2012/3/30/KAIST_08162012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of <i>0109</i></div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">There was no vector prepared for Moth 0109, so we did 5ml pre-culture on 10ml tube. Cell was grown about 16hours after pre-culture, and we did vector mini-prep on the sample. The concentration of the mini-prepped vector is 726.8ng/ul and the purity(A<sub>260</sub>/A<sub>280</sub>) was 1.86.</span><br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi electro-transformation into MG1655</div><br />
<br />
<div id="content_note" ><br />
<br />
<span>Strain: MG1655</br></br><br />
Plasmid source:</br></br><br />
<ul style="list-style-type:square"><br />
<li>pPoC: from DH5α-pPoC colony #1 and TOP10-pPoC colony #1</li><br />
<li>pPoCpi: from TOP10-pPoC colony #1 and #3</li><br />
</ul><br />
</br></br><br />
</span><br />
<br />
</div> <br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="17"><br />
<div class="date">August 17<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Expression check of Formate dehydrogenase.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0817Fig1" src="https://static.igem.org/mediawiki/2012/3/34/KAIST_08172012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 40.xx kDa protein, formate dehydrogenase. The band appeared more significantly than the case of pTrcHis2A vector. Therefore, using pBAD promoter seems to produce more proteins.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">PCR amplification – second trial</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0817Fig2" src="https://static.igem.org/mediawiki/2012/5/5b/KAIST_08172012_Fig2.png" /></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">As the band was not significant for <i>1203</i> and <i>1198</i> on previous trial and purity was low for <i>1191</i>, we tried secondary PCR amplification on samples for Gibson assembly; <i>1202</i>, <i>1203</i>, <i>1198</i>, <i>1191</i>. Particularly for <i>1202</i> gene, we tried additional colony PCR, so that we could check any problem with vector mini-prep. (PCR solution for <i>1202</i> colony PCR was : 22ul </span><br />
<br />
</br></br><br />
<div align="center"><img id="figure" alt="0817Fig3" src="https://static.igem.org/mediawiki/2012/e/e7/KAIST_08172012_Fig3.png" /></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</div><br />
<br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">MG1655-pPoC, MG1655-pPoCpi streaking</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">We could not distinguish single colony from plates. Therefore we scraped cells and streaked onto new plates.</span><br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">Wild-type <i>E.coli</i> showed more reddish and yellowish color than competent strains, DH5α and TOP10. The promoter BBa_J23119 may show different strength in-between strains.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">TOP10-pTrcHis2A_Bxb1 expression check </div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">TOP10 containing pTrcHis2A was cultured in 30mL of LB with 1% of Ampicillin.</br></span><br />
<span id="little">IPTG induced when OD = 0.4</br></span><br />
<span id="little">Sampling: 8 hours after induction, 1010 cells.</br></span><br />
<span id="little">Only soluble fractions were PAGEed.</br></span><br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">August 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:262px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">PCR amplification of <i>1202</i>, <i>0109</i> gene with original primer and Gibson primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/b0/KAIST_08182012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Each lane indicates:</br></br><br />
<ul><br />
<li><i>Moth_1202_1</i> : Gibson forward + original reverse</li><br />
<li><i>Moth_1202_2</i> : Gibson reverse + original forward</li><br />
<li><i>Moth_0109_1</i> : Gibson forward + original reverse</li><br />
<li><i>Moth_0109_2</i> : Gibson reverse + original forward</li><br />
</ul><br />
</br><br />
There was correct size and unspecific size of band on <i>1202_1</i> sample. Although there are also unspecific band, <i>1202_2</i> sample has large band of correct size. <i>0109_1</i> sample has only unspecific band and <i>0109_2</i> sample has right size of band. <br />
</span><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">When the PCR was unsuccessful, we first thought it might be problem of forward primer as in the case of <i>1199</i> and <i>1191</i>. Our guess was correct, and we’re going to fix the problem with forward primers. </span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1202</i> and <i>0109</i> forward primer design</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">We designed new forward primer hoping no problem with it. </span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/64/120818_1202_0109_primer_design.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Newly designed primers contain overlapping region with former DNA fragments, artificial ribosome binding site, and overlapping region with 1202 and 0109 each. </br></br><br />
</span><br />
<br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">MG1655-pPoC, MG1655-pPoCpi colony inoculation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Two colonies from each plate were inoculated into 3mL of LB containing 1% of chloramphenicol.</span><br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<br />
<span id="little">In 12 hours, cell cultures show colors compared to another common <i>E.coli</i> culture. When they are centrifuged, pellet shows clear and definite yellow or red color.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pTrcHis2A_Bxb1 electro-transformation into MG1655-pPoC, MG1655-pPoCpi</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pTrcHis2A_Bxb1 is electro-transformed into MG1655 containing pPoC and pPoCpi.</br></br><br />
Plasmid source: TOP10-pTrcHis2A_Bxb1</span><br />
</br></br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="19"><br />
<div class="date">August 19<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">August 20<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">August 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="22"><br />
<div class="date">August 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Primer design for Bxb1 integrase cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Primers for Bxb1 integrase cloning were designed. For controlling basal transcription level optimization, a mutant primer which has GTG rather than ATG for its start codon was also ordered.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0822Fig1" src="https://static.igem.org/mediawiki/2012/9/92/KAIST_2nd_08222012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">August 23<sup>rd</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="24"><br />
<div class="date">August 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title"> <i>0109</i>, <i>1191</i> PCR with new primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/1/10/120824_0109_1191_pcr_gel_picture.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">PCR product of Moth 0109 gene and 1191 gene has run on the gel. There was only faint band of 0109 fragment but the product band of 1191 gene was significantly shown on the gel. After gel electrophoresis, we tried PCR of 0109 once more and got significant amount of fragments. </span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">August 25<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC-Bxb1 double transformants inoculation for prep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Double transformants 1 through 24 were inoculated into 3ml of LB containing 1% of Cm and Ap. For our convenience, three colonies were inoculated in one culture tube.</span><br />
</br></br><br />
<br />
</div> <br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
<section id="26"><br />
<div class="date">August 26<sup>th</sup> 2012</div></br><br />
<br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC-Bxb1 double transformants plasmid DNA miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Inoculated cultures were mipi-preped. And Bxb1-pTrc plasmid was checked by PCR. In case of pPoC, it would be check by color.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<span id="little">Vector</span></br></br><br />
<div align="center"><img id="figure" alt="2_0826Fig1" src="https://static.igem.org/mediawiki/2012/3/38/KAIST_2nd_08252012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<span id="little">Confirm PCR</span></br></br><br />
<div align="center"><img id="figure" alt="2_0826Fig2" src="https://static.igem.org/mediawiki/2012/b/b9/KAIST_2nd_08252012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<span id="little">In confirm PCR, all the colonies seem to have Bxb1 vector. Colony #1, 4, 7, 10, 13, 14, 16, 19 and 22 will be inoculated.</span></br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1 PCR with new primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">For cloning of Bxb1 into pBAD and mutating its start codon from ATG to GTG, we PCR pGEM-B1-Bxb1 vector as template. Samples were duplicated</br></br></span><br />
<span><br />
PCR conditions<br />
<ol><br />
<li>95℃ for 3min</li><br />
<li>95℃ for 30sec</li><br />
<li>55℃ for 30sec</li><br />
<li>72℃ for 1min 30sec</li><br />
Step 2 to 4: 34 cycles<br />
<li>72℃ for 10min</li><br />
<li>12℃ forever</li><br />
</ol><br />
</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0826Fig3" src="https://static.igem.org/mediawiki/2012/6/61/KAIST_2nd_08262012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Duplicated samples were purified in one DNA binding column.</br></br>ATG: 297.9ng/uL (purity: 1.90)</br>GTG: purify failed. Sample lost</br></span><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">There were multiple bands in our PCR products. We will PCR again Bxb1-GTG with different conditions. Template vector may show larger bands.</span><br />
</br><br />
<br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pGEM-Bxb1 template vector gel electrophoresis</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">To check whether the undesired bands were template vector, we diluted template vector with same conditions of PCR, 50X, and electrophoresized.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0826Fig4" src="https://static.igem.org/mediawiki/2012/e/ea/KAIST_2nd_08262012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">The band was different from undesired bands from Bxb1 ATG, GTG PCR products.</span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Insert DNA (Bxb1 integrase) and vector (pTrc and pBAD) cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little"></span><br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
<section id="27"><br />
<div class="date">August 27<sup>th</sup> 2012</div></br><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1-pTrc and Bxb1-pBAD electrotransformation into MG1655</div><br />
<br />
<div id="content_note" ><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1 PCR with changed conditions</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">For cloning of Bxb1 into pBAD and mutating its start codon from ATG to GTG, we PCR pGEM-B1-Bxb1 vector as template. Samples were duplicated</span><br />
</br></br><br />
<div align="center"><br />
<table style="border:0px;background-color:transparent;"><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;"><b>PCR conditions #1</b></td> <td style="padding:5px 20px 5px 20px;"><b>PCR conditions #2</b></td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">1) 95℃ for 3min</td> <td style="padding:5px 20px 5px 20px;">1) 95℃ for 3min</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">2) 95℃ for 30sec</td> <td style="padding:5px 20px 5px 20px;">2) 95℃ for 30sec</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">3) 56.5℃ for 30sec</td> <td style="padding:5px 20px 5px 20px;">3) 58℃ for 30sec</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">4) 72℃ for 1min 30sec</br> (Step 2 to 4: 34 cycles)</td> <td style="padding:5px 20px 5px 20px;">4) 72℃ for 1min 30sec</br> (Step 2 to 4: 34 cycles)</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">5) 72℃ for 10min</td> <td style="padding:5px 20px 5px 20px;">5) 72℃ for 10min</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">6) 12℃ forever </td> <td style="padding:5px 20px 5px 20px;">6) 12℃ forever</td><br />
</tr><br />
<br />
</table><br />
<br />
</div><br />
</br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0827Fig1" src="https://static.igem.org/mediawiki/2012/a/a5/KAIST_2nd_08272012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">Still, undesired bands appeared. We do not know where they come from. We decide to extract desired DNA with gel extraction.</span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1, with GTG as start codon, PCR product gel extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">PCR products were gel extracted.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<ul style="list-style-type:square"><br />
<li>Bxb1 GTG cond. #1 - #1: 128.2ng/uL (purity: 1.89)</li><br />
<li>Bxb1 GTG cond. #1 - #2: 79.0ng/uL (purity: 1.89)</li><br />
<li>Bxb1 GTG cond. #2 - #1: 110.7ng/uL (purity: 1.89)</li><br />
<li>Bxb1 GTG cond. #2 - #2: 66.7ng/uL (purity: 1.94)</li><br />
</ul><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<span id="little"> Bxb1 GTG cond. #1 - #1 and Bxb1 GTG cond. #2 - #1 will be used to cloning.</span></br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Insert DNA (Bxb1 GTG) and vector (pTrc and pBAD) cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Insert DNA: Bxb1 integrase mutated by primer.</span></br><br />
<span id="little">Vectro DNA: pTrcHis2A, pBAD-mycHisC</span></br></br><br />
<span id="little">Prepared DNA concentration:</span></br></br><br />
<span><br />
<ul style='list-style-type:square'><br />
<li>Bxb1 GTG cond. #1 - #1: 128.2ng/uL (purity: 1.89)</li><br />
<li>Bxb1 GTG cond. #2 - #1: 110.7ng/uL (purity: 1.89)</li><br />
<li>pTrcHis2A: 116.1ng/uL</li><br />
<li>pBAD-mycHisC: 128.7ng/uL</li><br />
</ul><br />
</span><br />
</br></br><br />
<span id="little">Enzyme digestion: 37℃ for 1hr 30min, 80℃ for 20min</span></br></br> <br />
<br />
<div align="center"><img id="figure" alt="2_0827Fig2" src="https://static.igem.org/mediawiki/2012/c/c7/KAIST_2nd_08272012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<span id="little">Dephosphorylation: 37℃ for 30min, 70℃ for 5min</span></br></br><br />
<div align="center"><img id="figure" alt="2_0827Fig3" src="https://static.igem.org/mediawiki/2012/6/67/KAIST_2nd_08272012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<span id="little">Ligation: 16℃ overnight</span></br></br><br />
<div align="center"><img id="figure" alt="2_0827Fig4" src="https://static.igem.org/mediawiki/2012/8/8b/KAIST_2nd_08272012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1, pPoC double transformants colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Colony #1, 4, 7, 10, 13, 14, 16, 19 and 22 from master plate</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0827Fig5" src="https://static.igem.org/mediawiki/2012/4/45/KAIST_2nd_08272012_Fig5.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">The gel showed that all the colonies have Bxb1-pTrc vector. And also they all have pPoC or pPoCpi, because they show color.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="28"><br />
<div class="date">August 28<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1-GTG-pTrc and Bxb1-GTG-pBAD electrotransformation into MG1655</div><br />
<br />
<div id="content_note" ><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1-pTrc and Bxb1-pBAD transformants check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<span>Bxb1-pTrc: No colony</br><br />
Bxb1-pBAD: Colonies appeared and incoculated into 3mL of LB containing 1% of Ap</br><br />
</span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1-pTrc and Bxb1-pBAD transformants colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="2_0828Fig1" src="https://static.igem.org/mediawiki/2012/1/15/KAIST_2nd_08282012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1-pTrc and Bxb1-pBAD transformants plasmid DNA miniprep</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span>Colony #2: 109.8ng/uL, purity = 1.87</br><br />
Colony #3: 138.9ng/uL, purity = 1.74</br><br />
</span><br />
</br></br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">mRFP, GFP 30mL culture and sampling</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<section id="29"><br />
<div class="date">August 29<sup>th</sup> 2012</div></br><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 pBADmycHisC prepped plasmid DNA RE cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">mini-prepped plasmid DNAs were cut with SnaBI</br></br><br />
expected size : 5.6kb<br />
</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0829ig1" src="https://static.igem.org/mediawiki/2012/4/45/KAIST_2nd_08292012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"></span><br />
<br />
</br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title"> <i>1202</i> PCR to make fragments for Gibson assembly</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7c/120829.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">PCR for Moth 1202 gene has failed. No band for Moth 1202 gene has appeared on the gel.</span><br />
<br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Gibson assembly – assembly of pcr fragments.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Among three methods to assemble DNA fragments, we chose first method, which is assembly of PCR amplified DNA fragments. </br><br />
0.2umoles of each fragments are added into the reaction sample, and volume to be added for each fragments has calculated on the assumption that average MW of each base is 660Da. Table below is the result of calculation.</br><br />
Fragment 1</br><br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little">Fragment 2</br><br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/3/34/120829_table2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/98/120829-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">As we can only assemble the fragments through Gibson Assembly method and not amplify them, there would be very small amount of template remaining in the tube. We thought that’s why we couldn’t see any band on the gel. So we did PCR purification of the fragment once more and saw if the band appears on the gel. </span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="30"><br />
<div class="date">August 30<sup>th</sup> 2012</div></br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:262px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></br><br />
<div class="note-title"> Gibson Assembly product – Amplification by nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/67/120830.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">There was no right size of the band but just unspecific bands on the gel. <br />
There might be several reasons why the Gibson assembly ended with failure. So we decided to change the method of Gibson assembly; we will assemble fragments in the presence of vectors, so that assembly product can become stable, circular form. To cut the cost used in experiment, we will not design primers, but digest the vector with restriction enzymes to make it linear form.</span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Gibson primer second design 2</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/b7/120830table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">We designed primers again for another trial of Gibson Assembly. As stated before, We’ll use vector-involved assembly method to stabilize the assembly product and amplify them by in vivo system. </br>Like the method we used before, we’ll assemble 5 fragments first and assemble those 5-fragment-long two fragments into full construct. </br></span><br />
<br />
</div><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Double transformants expression SDS-PAGE</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Gel composition: 20%</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<span>pPoC</span><br><br><br />
<div align="center"><img id="figure" alt="2_0830Fig1" src="https://static.igem.org/mediawiki/2012/b/b6/KAIST_2nd_08302012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span>pPoCpo</span><br><br><br />
<div align="center"><img id="figure" alt="2_0830Fig2" src="https://static.igem.org/mediawiki/2012/a/a7/KAIST_2nd_08302012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Results</b></br></br><br />
<span id="little">We cannot see expression level change of fluorescent protein in SDS-PAGE.</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="31"><br />
<div class="date">August 31<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
</br></br><br />
</div><br />
</div><br />
<br />
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<br />
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<div style="clear:both;"></div><br />
<br />
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<br />
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<br />
<br />
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<br />
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<br />
<div id="content_note" ><br />
<br />
<span id="little">CONTENT2</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_08##Fig#" src="#"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
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<br />
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Mandie
http://2012.igem.org/File:120830table.PNG
File:120830table.PNG
2012-10-27T02:11:45Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:120830.PNG
File:120830.PNG
2012-10-27T02:07:17Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_10
Team:KAIST Korea/Notebook Labnote/2012 10
2012-10-27T02:04:36Z
<p>Mandie: </p>
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<h1>Labnote</h1><br />
<span id="sub-title">October</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">October 1<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
<section id="2"><br />
<div class="date">October 2<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">bFMO template preparation with new primer</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/a3/121002.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>We have found that our bFMO sequence was wrong. So that we have ordered new primers with right sequence.</br></span><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> OE PCR template preparation</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7f/121002_OE_pcr.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> pAutoIntegrase cloning check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/72/121002_pAutoIntegrase_cloning_check.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">We have checked the result of cloning with various ways but we can’t sure about the success of cloning.</span><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="3"><br />
<div class="date">October 3<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Visualization of inversion from GFP to RFP</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little"></br>bfmo gene is constructed in the vector(pJ401, Km registance) and transformed into DH5α.</br>50λ cell from stock is inoculated in 5mL LB </br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Visualization of inversion from GFP to RFP</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
<span id="little">We sampled the cell every 3hr. We induced one group of samples and un-induced another group of samples as a control. We took the picture of cell with confocal microscope at absorbance 500nm for GFP expression and 600nm for RFP expression. </br><br />
</span><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/ae/121003.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">We obtained proper data. Un-induced sample doesn’t show measurable RFP expression throughout the culture. However, induced sample shows the increase of RFP expression. Increase of GFP level in induced sample is due to the stability of GFP and time delay before inversion. </span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Insert DNA preparation with OE pcr</div><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/24/121003-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/87/121003-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8a/121003-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">October 4<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">October 5<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">Regional Jamboree at HKUST!!!!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">October 6<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">Regional Jamboree at HKUST!!!!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">October 7<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">Regional Jamboree at HKUST!!!!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="8"><br />
<div class="date">October 8<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">October 9<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">October 10<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">October 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">October 12<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">October 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">October 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">October 15<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">October 16<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">October 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="18"><br />
<div class="date">October 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/>Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Cell culture in 96-well plate for Indigo expression time optimization</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little"></br>We set the O.D value to 0.1 and cultured 1.5ml cell in 96 deep well plate. At each time point, we transferred 1ml cell from 96 deep well plate to EP tube and 200λ cell to flat 96 well plate for O.D check.</br><br />
After cell down in EP tube, we discarded the supernatant and resuspended the pellet in 200λ DMSO. We did sonication to the sample and after another cell down, we took the supernatant for TECAN measurement. Also, we checked absorbance after 3 days incubation and before 3days incubation.<br />
</br>We used TECAN at absorbance 620nm to measure the quantity of indigo production</br></br></span><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/3/3f/Tecan-1.PNG"></img></div><br />
<div style="clear:both;"></div></br><br />
<br />
<div align="center"><img id="figure" src="https://static.igem.org/mediawiki/2012/7/73/Tecan-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<span id="little"></br></br>We subtracted absorbance value of pure LB from value of samples and we set all samples to have same amount of cell using O.D value </br><br />
Graph shown below is indigo intensity per cell. After 3 days incubation, indigo seems to be degraded and all the samples seem to have almost same indigo level.</br></br></span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fc/Tecan_indigo-1.PNG"></img></div><br />
<div style="clear:both;"></div></br><br />
<br />
<span id="little"></br>We excluded 0hr data from the graph to see teh rising tendency clearly.</br></br></span><br />
<br />
<div align="center"><img id="figure" src="https://static.igem.org/mediawiki/2012/0/06/Tecan_indigo-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b></br>Discussion</b></br><br />
<br />
<span id="little"></br>This data shows that the cell successfully produces indigo. We can use this gene as a reporter of our module. However, we only checked the increase of indigo. It may decrease after prolonged culture. We should do additional experiments to get the peak in the graph and optimize the production time of indigo. </br></span><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a> <br />
<br />
</section><br />
<section id="19"><br />
<div class="date">October 19<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">October 20<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">October 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="22"><br />
<div class="date">October 22<sup>nd</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">October 23<sup>rd</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">October 24<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="25"><br />
<div class="date">October 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">October 26<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">October 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">October 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">October 29<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">October 30<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="31"><br />
<div class="date">October 31<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
</br></br><br />
</div><br />
</div><br />
<br />
</body><br />
</html><br />
{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:121002_pAutoIntegrase_cloning_check.PNG
File:121002 pAutoIntegrase cloning check.PNG
2012-10-27T02:03:19Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:121002_OE_pcr.PNG
File:121002 OE pcr.PNG
2012-10-27T02:01:23Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_8
Team:KAIST Korea/Notebook Labnote/2012 8
2012-10-27T01:54:32Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
{{:Team:KAIST_Korea/navi}}<br />
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margin-left:auto;<br />
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list-style-type:decimal;<br />
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{<br />
font-family: 'Open Sans', sans-serif;<br />
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padding:2px 10px 10px 10px;<br />
margin:0px 0px 35px 0px;<br />
background-image:url('https://static.igem.org/mediawiki/2012/a/a0/KAIST_labnote_logo.png');<br />
background-repeat:no-repeat;<br />
opacity:0.9;<br />
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}<br />
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</head><br />
<br />
<body><br />
<div id="main"><br />
<img id="top-img" src="https://static.igem.org/mediawiki/2012/6/6d/Back_home.PNG"/><br />
<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
</div><br />
<br />
<div id="top-img-description-box"><span id="top-img-description">Notebook : Labnote-August</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">August</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">August 1<sup>st</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">PCR amplification of <i>Moth_1191</i>, <i>1199</i> gene : decreasing annealing temperature down to 45℃</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">To make sure there’s no problem with unspecific binding or annealing temperature, we decreased annealing condition down to 45℃.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0801Fig1" src="https://static.igem.org/mediawiki/2012/e/e1/KAIST_08012012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">There was no band and it became certain that there are no problems with annealing procedure.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Induction of <i>Moth_0109</i> gene (sampling) and running SDS-PAGE gel. </div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">We induced <i>Moth_0109</i> gene in pBAD/myc vector with different Arabinose conditions and temperature: 10mM, 30mM and 37℃, 30℃. And then, we run the sample into SDS-PAGE gel. The gel is stained with coomassie blue for 1 hour and de-stained for overnight.</span><br />
</br></br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">PCR amplification of <i>Moth_1191</i>, <i>1199</i> gene with Gibson and TOPO primer</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">To see whether there are any problem with Gibson primers that we’ve designed, we did one more pcr amplification. Primers for Gibson assembly and TOPO cloning are mixed, and we could see any problems with forward or reverse primers. </span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0801Fig2" src="https://static.igem.org/mediawiki/2012/6/62/KAIST_08012012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Each lane indicates:</br></br><br />
<ul style="list-style-type:square"><br />
<li><i>Moth_1191_1</i> : TOPO forward primer and Gibson reverse primer</li> <br />
<li><i>Moth_1191_2</i> : Gibson forward primer and TOPO reverse primer</li><br />
<li><i>Moth_1199_1</i> : Gibson forward primer and TOPO reverse primer</li><br />
<li><i>Moth_1199_2</i> : TOPO forward primer and Gibson reverse primer</li><br />
</ul><br />
</br><br />
Correct size of band appeared when topo forward primer and Gibson reverse primer was used in amplification of both genes. Therefore, we can conclude that there is some unknown problem with forward primers.</span><br />
<br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi OE PCR with changed primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0801Fig1" src="https://static.igem.org/mediawiki/2012/8/81/KAIST_2nd_08012012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">→ Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="2"><br />
<div class="date">August 2<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Primer design for FDH derived from <i>Candida boidinii</i> mutant</div><br />
<br />
<div id="content_note" ><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0802Fig1" src="https://static.igem.org/mediawiki/2012/b/b8/KAIST_08022012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">These primers are universial primer that can be used for insertion of this gene into both pTrcHis2A vector and pBAD/mycHisC vector. </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Expression check of <i>Moth_0109</i> gene on pBAD/mycHisC vector (Result)</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 59.94 kDa protein, formate tetrahydrofolate ligase. </span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0802Fig2" src="https://static.igem.org/mediawiki/2012/7/7c/KAIST_08022012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">The band of PAGE gel was not certain, so we </span><br />
<!--이것도 쓰다가 말았음;; --><br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi cloning - Digestion</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Restriction enzyme digestion</span><br />
</br></br><br />
<span><br />
<ul style="padding:0px 0px 0px 40px;"><br />
<li>Enzyme pre-mix / each reaction – 37℃ 1hr, 80℃ 20min</li></br><br />
<div align="center"><img id="figure" alt="2_0802Fig1" src="https://static.igem.org/mediawiki/2012/1/1e/KAIST_2nd_08022012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<li>Digestion mixture – 37℃ 1hr, 80℃ 20min</li></br><br />
<div align="center"><img id="figure" alt="2_0802Fig2" src="https://static.igem.org/mediawiki/2012/7/79/KAIST_2nd_08022012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</ul><br />
</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1 integrase transformation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Synthesized gene arrived.</br></span><br />
<span id="little">Transformed into DH5α </span><br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<section id="3"><br />
<div class="date">August 3<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 integrase-DH5α streaking</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">Synthesized gene was cloned into pGEM-B1 vector. It seems the vector concentration is too high so that too much colonies may appeared.</span><br />
<br />
</br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi cloning - Ligation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Ligation - 16℃ overnight</span><br />
</br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0803Fig3" src="https://static.igem.org/mediawiki/2012/8/81/KAIST_2nd_08032012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Transformed into DH5α chemically competent cell.</span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">BBa_J04450 transformation into DH5α</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">For pSB1C3 vector amplification</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<section id="4"><br />
<div class="date">August 4<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title"><i>1191</i> and <i>1199</i> forward primer design</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">We designed new forward primers for both <i>1191</i> and <i>1199</i> gene that are thought to solve problems appeared during PCR amplification.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0804Fig1" src="https://static.igem.org/mediawiki/2012/5/59/KAIST_08042012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_J04450 and Bxb1-pGEM colony picking and inoculation.</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Made master plates and inoculated in 3mL of LB.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="5"><br />
<div class="date">August 5<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="6"><br />
<div class="date">August 6<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_J04450 Plasmid miniprep</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_J04450(pSB1C3): 285.8ng/uL</span></br><br />
<span id="little">purity:2.10</span><br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Requested part arrived</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Each parts were streaked on appropriate plates.</br></br><br />
<ul><br />
<li>BBa_I11020</li><br />
<li>BBa_I11022</li><br />
<li>BBa_I11023</li><br />
<li>BBa_I11030</li><br />
<li>BBa_K137007</li><br />
<li>BBa_K137008</li><br />
<li>BBa_K137010</li><br />
<li>BBa_K112001</li><br />
<li>BBa_K112141</li><br />
<li>BBa_K112142</li><br />
</ul><br />
</span><br />
</br></br><br />
<br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">BBa_K381001(GFP) and BBa_K398005(pSB1C3) transformation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">BBa_K381001(GFP): for GFP control</span></br><br />
<span id='little'>BBa_K398005(pSB1C3): for pSB1C3 template</span></br><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="7"><br />
<div class="date">August 7<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">FDH gene PCR amplification</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0807Fig1" src="https://static.igem.org/mediawiki/2012/7/7b/KAIST_08072012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">We amplified 2 samples of formate dehydrogenase gene with the primers we designed. From gel electrophoresis picture, we can say that oligonucleotides with the proper size 1100bp are present in the sample.</span><br />
<div align="center"><img id="figure" alt="0807Fig2" src="https://static.igem.org/mediawiki/2012/e/eb/KAIST_08072012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br> <br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">BBa_J04450(pSB1C3) enzyme cut check(single cut with EcoRI)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0807Fig1" src="https://static.igem.org/mediawiki/2012/9/93/KAIST_2nd_08072012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Vector was double digested. The insert and vector sizes were correct</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi OE PCR product nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0807Fig2" src="https://static.igem.org/mediawiki/2012/9/91/KAIST_2nd_08072012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">→ Gel extracted</span><br />
<br />
</br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">BBa_K398005 Plasmid miniprep & single cut check with EcoRI</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0807Fig3" src="https://static.igem.org/mediawiki/2012/c/c9/KAIST_2nd_08072012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pPoC insert</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0807Fig4" src="https://static.igem.org/mediawiki/2012/d/d5/KAIST_2nd_08072012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"><br />
<ul style="list-style-type:square; padding:0px 0px 0px 40px;"><br />
<li>HSTaq Pol MM : 25uL</li><br />
<li>DW : 1uL</li><br />
<li>templates : 22uL</li><br />
<li>primers : 2uL</li><br />
<br />
</ul><br />
</span></br></br><br />
<br />
<span id="little">pPoCpi insert</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="2_0807Fig4" src="https://static.igem.org/mediawiki/2012/d/d5/KAIST_2nd_08072012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"><br />
<ul style="list-style-type:square; padding:0px 0px 0px 40px;"><br />
<li>HSTaq Pol MM : 25uL</li><br />
<li>templates : 23.2uL</li><br />
<li>primers : 2uL</li><br />
</ul><br />
</span><br />
</br><br />
</br><br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="2_0807Fig5" src="https://static.igem.org/mediawiki/2012/e/ea/KAIST_2nd_08072012_Fig5.png"></img></div><br />
<div style="clear:both;"></div><br />
</br><br />
<span id ="little">→ Concentration after gel extraction was not sufficient for cloning</span></br></br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping PCR template preparation</div><br />
<br />
<div id="content_note" ><br />
<br />
<div align="center"><img id="figure" alt="2_0807Fig6" src="https://static.igem.org/mediawiki/2012/9/99/KAIST_2nd_08072012_Fig6.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<div align="center"><img id="figure" alt="2_0807Fig7" src="https://static.igem.org/mediawiki/2012/3/3a/KAIST_2nd_08072012_Fig7.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<br />
<br />
<section id="8"><br />
<div class="date">August 8<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">FDH gene and pTrcHis2A, pBAD/mycHisC vector enzyme digestion</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We mixed restriction enzymes and buffers with DNA samples as shown below.</span></br></br><br />
<br />
<div align="center"><img id="figure" alt="0808Fig1" src="https://static.igem.org/mediawiki/2012/3/31/KAIST_08082012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<span id="little">After mixing the enzymes and buffer with the sample, we incubated the sample on 37℃ for 1 hour and then on 65℃ for 20minutes. </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pTrcHis2A, pBAD/myc vector dephosphorylation</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<span id="little">We mixed phosphatidase and buffer with DNA sample as shown below.</span></br></br><br />
<div align="center"><img id="figure" alt="0808Fig2" src="https://static.igem.org/mediawiki/2012/4/49/KAIST_08082012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">After mixing the enzyme and buffer with the sample, we incubated the sample on 37℃ for 1 hour 20minutes and then on 70℃ for 5minutes. </span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">ligation of FDH gene with pTrcHis2A and pBAD/mycHisC vectors</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<span id="little">We mixed ligase and buffer with DNA sample as shown below.</span><br />
</br></br><br />
<div align="center"><img id="figure" alt="0808Fig3" src="https://static.igem.org/mediawiki/2012/4/4e/KAIST_08082012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">After mixing the enzyme and buffer with the sample, we incubated the sample on 16℃ for 16 hours.</span><br />
<br />
</br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Primer amplification of <i>1191</i> and <i>1199</i></div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0808Fig4" src="https://static.igem.org/mediawiki/2012/5/5d/KAIST_08082012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pPoC insert</span></br></br><br />
<div align="center"><img id="figure" alt="2_0808Fig2" src="https://static.igem.org/mediawiki/2012/9/9b/KAIST_2nd_08082012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">pPoCpi insert</span></br></br><br />
<div align="center"><img id="figure" alt="2_0808Fig3" src="https://static.igem.org/mediawiki/2012/5/53/KAIST_2nd_08082012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<span><br />
<ul style="list-style-type:square"><br />
<br />
<li>HSTaq Pol MM : 25uL</li><br />
<li>DW : 5.2uL</li><br />
<li>templates : 19.8uL</li><br />
<li>primers : 2uL</li><br />
<li>pfu-x : 0.1uL</li><br />
<br />
</ul><br />
</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0808Fig1" src="https://static.igem.org/mediawiki/2012/3/31/KAIST_2nd_08082012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">→ Gel extraction</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping PCR template preparation</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0808Fig4" src="https://static.igem.org/mediawiki/2012/5/5d/KAIST_2nd_08082012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<br />
<section id="9"><br />
<div class="date">August 9<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">vector transformation into TOP10 cell</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Transformation of pTrcHis2A vector into TOP10 cell was successful. So we did vector mini-preparation after the transformation. However, transformation of pBAD vector has failed, so we ligated the samples again. </span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0809Fig1" src="https://static.igem.org/mediawiki/2012/e/e9/KAIST_08092012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">This is gel picture to confirm that FDH is well inserted into TOP10 cells. 3 wells on the left are PCR product of <!--??/--></span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 integrase cloning into pTrcHis2a and pBADmycHisC</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Bxb1 integrase: 1515bp</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0809Fig1" src="https://static.igem.org/mediawiki/2012/3/33/KAIST_2nd_08092012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">→ Gel extracted</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Extracted OE PCR product cloning into pSB1C3</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pSB1C3 was originated from BBa_K398005.</br></br>Transformed into TOP10</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<span id="little">Transformed colony was grown in LB media. No colony shows fluorescence. </span><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">We think cloning failed.</span><br />
<br />
</br></br><br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="10"><br />
<div class="date">August 10<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title"> Bxb1-pTrc, pBAD colony inoculation</div><br />
<br />
<div id="content_note" ><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Overlapping PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">In this time, PCR was done first 18 cycles without any primer.</br><br />
After that, we add 1uL of primers. And then PCR was done 20 cycles more.<br />
</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0810Fig1" src="https://static.igem.org/mediawiki/2012/7/79/KAIST_2nd_08102012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">With this OE PCR conditions, yield was better than previous conditions.</span><br />
<br />
</br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pPoC, pPoCpi insert cloning into pSB1C3</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pSB1C3 was derived from BBa_K398005<br />
</span><br />
</br></br><br />
<br />
<span><br />
<ol style="padding:0px 0px 0px 40px;"><br />
<li>Digestion – 37℃ 1hr, 80℃ 20min</li><br />
<div align="center"><img id="figure" alt="2_0810Fig5" src="https://static.igem.org/mediawiki/2012/5/5d/KAIST_2nd_08102012_Fig5.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<li>Dephosphorylation – 37℃ 30min, 80℃ 10min</li><br />
<div align="center"><img id="figure" alt="2_0810Fig6" src="https://static.igem.org/mediawiki/2012/8/87/KAIST_2nd_08102012_Fig6.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<li>Ligation – 37℃ 2hr</li><br />
<div align="center"><img id="figure" alt="2_0810Fig7" src="https://static.igem.org/mediawiki/2012/f/fb/KAIST_2nd_08102012_Fig7.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</ol><br />
</span><br />
</br></br><br />
<span id="little">Ligated products were transformed into DH5α.</span><br />
<br />
<br />
</br><br />
</div><br />
</br><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Transformed cell inoculation and colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0810Fig8" src="https://static.igem.org/mediawiki/2012/3/3c/KAIST_2nd_08102012_Fig8.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">only pPoCpi construct cloning was completed </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="11"><br />
<div class="date">August 11<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">pTrcHis2A vector transformation into <i>E.coli</i> MG1655 cell</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pBAD/mycHisC-FDH transformation into TOP10</div><br />
<br />
<div id="content_note" ><br />
<br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 pTrcHis2A, pBADmycHisC plasmid DNA mini-prep</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">single cut check</span></br></br><br />
<div align="center"><img id="figure" alt="2_0811Fig1" src="https://static.igem.org/mediawiki/2012/d/db/KAIST_2nd_08112012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
<br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi nested PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<span id="little">nested PCR to prepare more insert DNA</span></br></br><br />
<div align="center"><img id="figure" alt="2_0811Fig2" src="https://static.igem.org/mediawiki/2012/1/11/KAIST_2nd_08112012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<br />
<section id="12"><br />
<div class="date">August 12<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">colony PCR check of pTrcHis2A inserted FDH gene</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0812Fig1" src="https://static.igem.org/mediawiki/2012/8/80/KAIST_08122012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">We checked the presence of FDH gene by conducting colony PCR. Correct size of band appeared at about 1115bp, which means formate dehydrogenase is inserted correctly. </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pBAD FDH MG1655 electroporation</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<br />
<span id="little">Fail</span><br />
<!--여기 쓸말 더 있으면 써줘요 지환오빠 ㅋㅋ--><br />
</br><br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="13"><br />
<div class="date">August 13<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">FDH pBAD MG1655 electroporation</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cells on 7:10 AM and did cell-down on 10:00AM. And then we plated cells on 1:00PM and checked colony on 8/14 5:00AM</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<!-- <br />
<div align="center"><img id="figure" alt="08##Fig#" src="#"></img></div><br />
<div style="clear:both;"></div> Colony pic.--><br />
</br><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">FDH pTrc MG1655 expression check (sampling and induction)</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cells (500λ of cells put into 50mL LB with 50λ Ampicillin) on 7 : 30 AM and checked Optical Density on 10:30pm. According to the O.D. value, we induced 37℃ sample on 10:30PM and did sampling on 4:30pm. Also, we induced 30℃ sample on 11:30pm and did sampling on 5:30pm. (6 hour induction) Each temperature samples were induced by different conditions ; 2mM and 3mM of IPTG.</span><br />
</br></br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoCpi cloning into pSB1C3</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Cloning was done by remaining DNA samples. </span><br />
</br></br><br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="14"><br />
<div class="date">August 14<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">colony PCR check of pBADmycHisC inserted FDH gene</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0814Fig1" src="https://static.igem.org/mediawiki/2012/e/e2/KAIST_08142012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">We checked the presence of FDH gene by conducting colony PCR. Correct size of band appeared at about 1115 bp, which means formate dehydrogenase is inserted correctly. </span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Expression check of Formate dehydrogenase on pTrcHis2A vector (Result)</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0814Fig2" src="https://static.igem.org/mediawiki/2012/6/64/KAIST_08142012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 43.7 kDa protein, formate dehydrogenase. The band appeared significantly on the correct size. </span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title">FDH pBAD MG1655 seeding for expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cell on 7:00PM, so we’ll incubate cell about 16hours. (This means we have to do next seeding on 11:00AM. )</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Primer design for the insertion of <i>1202</i> and <i>0109</i></div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0814Fig3" src="https://static.igem.org/mediawiki/2012/b/bd/KAIST_08142012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">To eliminate uncertain spacer between genes, we inserted </span><br />
<!--쓰다가 말았네;;--><br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 pTrcHis2A and pPoCpi ligated products transformation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">The products were transformed into TOP10</span><br />
</br></br><br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="15"><br />
<div class="date">August 15<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">FDH pBAD MG1655 expression check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little">We seeded cells on 11 : 30 AM and checked Optical Density on 1:30pm. According to the O.D. value, we induced 37℃ sample on 1:40pm and did sampling on 7:40pm. Also, we induced 30℃ sample on 3:30pm and did sampling on 9:30pm. (6 hour induction) Each temperature samples were induced by different conditions: 10mM of arabinose and 30mM of arabinose.</span><br />
<br />
</br><br />
<br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoCpi colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_08##Fig#" src="https://static.igem.org/mediawiki/2012/c/cd/KAIST_2nd_08152012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Among colonies #1~10, four colonies were inoculated.<br />
#1, 3, 4, 7<br />
</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1 pTrcHis2A colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0815Fig2" src="https://static.igem.org/mediawiki/2012/9/9d/KAIST_2nd_08152012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Among colonies #1~20, three colonies were inoculated.<br />
#2, 5, 19<br />
</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<section id="16"><br />
<div class="date">August 16<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">PCR amplification of <i>1201</i>, <i>1202</i>, <i>1203</i>, <i>1198</i> and <i>1191</i></div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0816Fig1" src="https://static.igem.org/mediawiki/2012/8/86/KAIST_08152012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">Primers arrived, and we began to do additional PCR amplification on the preparation for Gibson assembly. Each PCR sample has the correct size of oligonucleotides except <i>Moth 1202</i> gene.</br></br><br />
<ul><br />
<li><i>1191</i> : 921bp</br></li><br />
<li><i>1198</i> : 972bp + about 400bp = 1300 bp </br></li><br />
<li><i>1201</i> : 1341bp + ?</br></li><br />
<li><i>1202</i> : 2190bp + </br></li><br />
<li><i>1203</i> : 2025bp + 1600bp = 3625bp</br></li><br />
</ul><br />
</br><br />
We did PCR purification on the PCR product and the concentration/purity result was as shown below. <br />
</span><br />
</br></br><br />
<br />
<div align="center"><img id="figure" alt="0816Fig2" src="https://static.igem.org/mediawiki/2012/3/30/KAIST_08162012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Pre-culture of <i>0109</i></div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">There was no vector prepared for Moth 0109, so we did 5ml pre-culture on 10ml tube. Cell was grown about 16hours after pre-culture, and we did vector mini-prep on the sample. The concentration of the mini-prepped vector is 726.8ng/ul and the purity(A<sub>260</sub>/A<sub>280</sub>) was 1.86.</span><br />
</br><br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC and pPoCpi electro-transformation into MG1655</div><br />
<br />
<div id="content_note" ><br />
<br />
<span>Strain: MG1655</br></br><br />
Plasmid source:</br></br><br />
<ul style="list-style-type:square"><br />
<li>pPoC: from DH5α-pPoC colony #1 and TOP10-pPoC colony #1</li><br />
<li>pPoCpi: from TOP10-pPoC colony #1 and #3</li><br />
</ul><br />
</br></br><br />
</span><br />
<br />
</div> <br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="17"><br />
<div class="date">August 17<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Expression check of Formate dehydrogenase.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0817Fig1" src="https://static.igem.org/mediawiki/2012/3/34/KAIST_08172012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">For each samples of different induction conditions, we run SDS-PAGE gel and checked the presence of 40.xx kDa protein, formate dehydrogenase. The band appeared more significantly than the case of pTrcHis2A vector. Therefore, using pBAD promoter seems to produce more proteins.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">PCR amplification – second trial</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0817Fig2" src="https://static.igem.org/mediawiki/2012/5/5b/KAIST_08172012_Fig2.png" /></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Discussion</b></br></br><br />
<span id="little">As the band was not significant for <i>1203</i> and <i>1198</i> on previous trial and purity was low for <i>1191</i>, we tried secondary PCR amplification on samples for Gibson assembly; <i>1202</i>, <i>1203</i>, <i>1198</i>, <i>1191</i>. Particularly for <i>1202</i> gene, we tried additional colony PCR, so that we could check any problem with vector mini-prep. (PCR solution for <i>1202</i> colony PCR was : 22ul </span><br />
<br />
</br></br><br />
<div align="center"><img id="figure" alt="0817Fig3" src="https://static.igem.org/mediawiki/2012/e/e7/KAIST_08172012_Fig3.png" /></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</div><br />
<br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">MG1655-pPoC, MG1655-pPoCpi streaking</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">We could not distinguish single colony from plates. Therefore we scraped cells and streaked onto new plates.</span><br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">Wild-type <i>E.coli</i> showed more reddish and yellowish color than competent strains, DH5α and TOP10. The promoter BBa_J23119 may show different strength in-between strains.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">TOP10-pTrcHis2A_Bxb1 expression check </div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">TOP10 containing pTrcHis2A was cultured in 30mL of LB with 1% of Ampicillin.</br></span><br />
<span id="little">IPTG induced when OD = 0.4</br></span><br />
<span id="little">Sampling: 8 hours after induction, 1010 cells.</br></span><br />
<span id="little">Only soluble fractions were PAGEed.</br></span><br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">August 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">PCR amplification of <i>1202</i>, <i>0109</i> gene with original primer and Gibson primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/b/b0/KAIST_08182012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Each lane indicates:</br></br><br />
<ul><br />
<li><i>Moth_1202_1</i> : Gibson forward + original reverse</li><br />
<li><i>Moth_1202_2</i> : Gibson reverse + original forward</li><br />
<li><i>Moth_0109_1</i> : Gibson forward + original reverse</li><br />
<li><i>Moth_0109_2</i> : Gibson reverse + original forward</li><br />
</ul><br />
</br><br />
There was correct size and unspecific size of band on <i>1202_1</i> sample. Although there are also unspecific band, <i>1202_2</i> sample has large band of correct size. <i>0109_1</i> sample has only unspecific band and <i>0109_2</i> sample has right size of band. <br />
</span><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">When the PCR was unsuccessful, we first thought it might be problem of forward primer as in the case of <i>1199</i> and <i>1191</i>. Our guess was correct, and we’re going to fix the problem with forward primers. </span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title"><i>1202</i> and <i>0109</i> forward primer design</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">We designed new forward primer hoping no problem with it. </span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/6/64/120818_1202_0109_primer_design.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Newly designed primers contain overlapping region with former DNA fragments, artificial ribosome binding site, and overlapping region with 1202 and 0109 each. </br></br><br />
</span><br />
<br />
</div><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">MG1655-pPoC, MG1655-pPoCpi colony inoculation</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Two colonies from each plate were inoculated into 3mL of LB containing 1% of chloramphenicol.</span><br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<br />
<span id="little">In 12 hours, cell cultures show colors compared to another common <i>E.coli</i> culture. When they are centrifuged, pellet shows clear and definite yellow or red color.</span><br />
<br />
</br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pTrcHis2A_Bxb1 electro-transformation into MG1655-pPoC, MG1655-pPoCpi</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">pTrcHis2A_Bxb1 is electro-transformed into MG1655 containing pPoC and pPoCpi.</br></br><br />
Plasmid source: TOP10-pTrcHis2A_Bxb1</span><br />
</br></br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="19"><br />
<div class="date">August 19<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="20"><br />
<div class="date">August 20<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="21"><br />
<div class="date">August 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="22"><br />
<div class="date">August 22<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Primer design for Bxb1 integrase cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Primers for Bxb1 integrase cloning were designed. For controlling basal transcription level optimization, a mutant primer which has GTG rather than ATG for its start codon was also ordered.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0822Fig1" src="https://static.igem.org/mediawiki/2012/9/92/KAIST_2nd_08222012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="23"><br />
<div class="date">August 23<sup>rd</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="24"><br />
<div class="date">August 24<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title"> <i>0109</i>, <i>1191</i> PCR with new primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/1/10/120824_0109_1191_pcr_gel_picture.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">PCR product of Moth 0109 gene and 1191 gene has run on the gel. There was only faint band of 0109 fragment but the product band of 1191 gene was significantly shown on the gel. After gel electrophoresis, we tried PCR of 0109 once more and got significant amount of fragments. </span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="25"><br />
<div class="date">August 25<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC-Bxb1 double transformants inoculation for prep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Double transformants 1 through 24 were inoculated into 3ml of LB containing 1% of Cm and Ap. For our convenience, three colonies were inoculated in one culture tube.</span><br />
</br></br><br />
<br />
</div> <br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
<section id="26"><br />
<div class="date">August 26<sup>th</sup> 2012</div></br><br />
<br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">pPoC-Bxb1 double transformants plasmid DNA miniprep & PCR check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Inoculated cultures were mipi-preped. And Bxb1-pTrc plasmid was checked by PCR. In case of pPoC, it would be check by color.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<span id="little">Vector</span></br></br><br />
<div align="center"><img id="figure" alt="2_0826Fig1" src="https://static.igem.org/mediawiki/2012/3/38/KAIST_2nd_08252012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<span id="little">Confirm PCR</span></br></br><br />
<div align="center"><img id="figure" alt="2_0826Fig2" src="https://static.igem.org/mediawiki/2012/b/b9/KAIST_2nd_08252012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<span id="little">In confirm PCR, all the colonies seem to have Bxb1 vector. Colony #1, 4, 7, 10, 13, 14, 16, 19 and 22 will be inoculated.</span></br><br />
<br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1 PCR with new primers</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">For cloning of Bxb1 into pBAD and mutating its start codon from ATG to GTG, we PCR pGEM-B1-Bxb1 vector as template. Samples were duplicated</br></br></span><br />
<span><br />
PCR conditions<br />
<ol><br />
<li>95℃ for 3min</li><br />
<li>95℃ for 30sec</li><br />
<li>55℃ for 30sec</li><br />
<li>72℃ for 1min 30sec</li><br />
Step 2 to 4: 34 cycles<br />
<li>72℃ for 10min</li><br />
<li>12℃ forever</li><br />
</ol><br />
</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0826Fig3" src="https://static.igem.org/mediawiki/2012/6/61/KAIST_2nd_08262012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little">Duplicated samples were purified in one DNA binding column.</br></br>ATG: 297.9ng/uL (purity: 1.90)</br>GTG: purify failed. Sample lost</br></span><br />
<br />
</br><br />
<b>Discussions</b></br></br><br />
<span id="little">There were multiple bands in our PCR products. We will PCR again Bxb1-GTG with different conditions. Template vector may show larger bands.</span><br />
</br><br />
<br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">pGEM-Bxb1 template vector gel electrophoresis</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">To check whether the undesired bands were template vector, we diluted template vector with same conditions of PCR, 50X, and electrophoresized.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0826Fig4" src="https://static.igem.org/mediawiki/2012/e/ea/KAIST_2nd_08262012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<br />
<span id="little">The band was different from undesired bands from Bxb1 ATG, GTG PCR products.</span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Insert DNA (Bxb1 integrase) and vector (pTrc and pBAD) cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little"></span><br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
<section id="27"><br />
<div class="date">August 27<sup>th</sup> 2012</div></br><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1-pTrc and Bxb1-pBAD electrotransformation into MG1655</div><br />
<br />
<div id="content_note" ><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1 PCR with changed conditions</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">For cloning of Bxb1 into pBAD and mutating its start codon from ATG to GTG, we PCR pGEM-B1-Bxb1 vector as template. Samples were duplicated</span><br />
</br></br><br />
<div align="center"><br />
<table style="border:0px;background-color:transparent;"><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;"><b>PCR conditions #1</b></td> <td style="padding:5px 20px 5px 20px;"><b>PCR conditions #2</b></td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">1) 95℃ for 3min</td> <td style="padding:5px 20px 5px 20px;">1) 95℃ for 3min</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">2) 95℃ for 30sec</td> <td style="padding:5px 20px 5px 20px;">2) 95℃ for 30sec</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">3) 56.5℃ for 30sec</td> <td style="padding:5px 20px 5px 20px;">3) 58℃ for 30sec</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">4) 72℃ for 1min 30sec</br> (Step 2 to 4: 34 cycles)</td> <td style="padding:5px 20px 5px 20px;">4) 72℃ for 1min 30sec</br> (Step 2 to 4: 34 cycles)</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">5) 72℃ for 10min</td> <td style="padding:5px 20px 5px 20px;">5) 72℃ for 10min</td><br />
</tr><br />
<tr><br />
<td style="padding:5px 20px 5px 20px;">6) 12℃ forever </td> <td style="padding:5px 20px 5px 20px;">6) 12℃ forever</td><br />
</tr><br />
<br />
</table><br />
<br />
</div><br />
</br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0827Fig1" src="https://static.igem.org/mediawiki/2012/a/a5/KAIST_2nd_08272012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">Still, undesired bands appeared. We do not know where they come from. We decide to extract desired DNA with gel extraction.</span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1, with GTG as start codon, PCR product gel extraction</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">PCR products were gel extracted.</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<ul style="list-style-type:square"><br />
<li>Bxb1 GTG cond. #1 - #1: 128.2ng/uL (purity: 1.89)</li><br />
<li>Bxb1 GTG cond. #1 - #2: 79.0ng/uL (purity: 1.89)</li><br />
<li>Bxb1 GTG cond. #2 - #1: 110.7ng/uL (purity: 1.89)</li><br />
<li>Bxb1 GTG cond. #2 - #2: 66.7ng/uL (purity: 1.94)</li><br />
</ul><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<span id="little"> Bxb1 GTG cond. #1 - #1 and Bxb1 GTG cond. #2 - #1 will be used to cloning.</span></br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Insert DNA (Bxb1 GTG) and vector (pTrc and pBAD) cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Insert DNA: Bxb1 integrase mutated by primer.</span></br><br />
<span id="little">Vectro DNA: pTrcHis2A, pBAD-mycHisC</span></br></br><br />
<span id="little">Prepared DNA concentration:</span></br></br><br />
<span><br />
<ul style='list-style-type:square'><br />
<li>Bxb1 GTG cond. #1 - #1: 128.2ng/uL (purity: 1.89)</li><br />
<li>Bxb1 GTG cond. #2 - #1: 110.7ng/uL (purity: 1.89)</li><br />
<li>pTrcHis2A: 116.1ng/uL</li><br />
<li>pBAD-mycHisC: 128.7ng/uL</li><br />
</ul><br />
</span><br />
</br></br><br />
<span id="little">Enzyme digestion: 37℃ for 1hr 30min, 80℃ for 20min</span></br></br> <br />
<br />
<div align="center"><img id="figure" alt="2_0827Fig2" src="https://static.igem.org/mediawiki/2012/c/c7/KAIST_2nd_08272012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
<span id="little">Dephosphorylation: 37℃ for 30min, 70℃ for 5min</span></br></br><br />
<div align="center"><img id="figure" alt="2_0827Fig3" src="https://static.igem.org/mediawiki/2012/6/67/KAIST_2nd_08272012_Fig3.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<span id="little">Ligation: 16℃ overnight</span></br></br><br />
<div align="center"><img id="figure" alt="2_0827Fig4" src="https://static.igem.org/mediawiki/2012/8/8b/KAIST_2nd_08272012_Fig4.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1, pPoC double transformants colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Colony #1, 4, 7, 10, 13, 14, 16, 19 and 22 from master plate</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0827Fig5" src="https://static.igem.org/mediawiki/2012/4/45/KAIST_2nd_08272012_Fig5.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussions</b></br></br><br />
<br />
<span id="little">The gel showed that all the colonies have Bxb1-pTrc vector. And also they all have pPoC or pPoCpi, because they show color.</span><br />
<br />
</br><br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="28"><br />
<div class="date">August 28<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1-GTG-pTrc and Bxb1-GTG-pBAD electrotransformation into MG1655</div><br />
<br />
<div id="content_note" ><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1-pTrc and Bxb1-pBAD transformants check</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<span>Bxb1-pTrc: No colony</br><br />
Bxb1-pBAD: Colonies appeared and incoculated into 3mL of LB containing 1% of Ap</br><br />
</span><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1-pTrc and Bxb1-pBAD transformants colony PCR</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<div align="center"><img id="figure" alt="2_0828Fig1" src="https://static.igem.org/mediawiki/2012/1/15/KAIST_2nd_08282012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<br />
</br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">Bxb1-pTrc and Bxb1-pBAD transformants plasmid DNA miniprep</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span>Colony #2: 109.8ng/uL, purity = 1.87</br><br />
Colony #3: 138.9ng/uL, purity = 1.74</br><br />
</span><br />
</br></br><br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
<div class="note-title">mRFP, GFP 30mL culture and sampling</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<br />
<section id="29"><br />
<div class="date">August 29<sup>th</sup> 2012</div></br><br />
</br><div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Bxb1 pBADmycHisC prepped plasmid DNA RE cut check</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">mini-prepped plasmid DNAs were cut with SnaBI</br></br><br />
expected size : 5.6kb<br />
</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_0829ig1" src="https://static.igem.org/mediawiki/2012/4/45/KAIST_2nd_08292012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"></span><br />
<br />
</br><br />
<img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:35px;float:bottom;"id="packman" src='https://static.igem.org/mediawiki/2012/0/03/KAIST_PACKMAN.png'/> PACKMAN<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title"> <i>1202</i> PCR to make fragments for Gibson assembly</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/7/7c/120829.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">PCR for Moth 1202 gene has failed. No band for Moth 1202 gene has appeared on the gel.</span><br />
<br />
</div><br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Gibson assembly – assembly of pcr fragments.</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<span id="little">Among three methods to assemble DNA fragments, we chose first method, which is assembly of PCR amplified DNA fragments. </br><br />
0.2umoles of each fragments are added into the reaction sample, and volume to be added for each fragments has calculated on the assumption that average MW of each base is 660Da. Table below is the result of calculation.</br><br />
</br>Fragment 1</br><br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/d/d8/120829_table.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>Fragment 2</br><br />
</span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/3/34/120829_table2.PNG"></img></div><br />
<div style="clear:both;"></div></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/9/98/120829-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">As we can only assemble the fragments through Gibson Assembly method and not amplify them, there would be very small amount of template remaining in the tube. We thought that’s why we couldn’t see any band on the gel. So we did PCR purification of the fragment once more and saw if the band appears on the gel. </span><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="30"><br />
<div class="date">August 30<sup>th</sup> 2012</div></br><br />
<br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:260px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:260px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<br />
<div class="note-title">Double transformants expression SDS-PAGE</div><br />
<br />
<div id="content_note" ><br />
<br />
<span id="little">Gel composition: 20%</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<span>pPoC</span><br><br><br />
<div align="center"><img id="figure" alt="2_0830Fig1" src="https://static.igem.org/mediawiki/2012/b/b6/KAIST_2nd_08302012_Fig1.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span>pPoCpo</span><br><br><br />
<div align="center"><img id="figure" alt="2_0830Fig2" src="https://static.igem.org/mediawiki/2012/a/a7/KAIST_2nd_08302012_Fig2.png"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<b>Results</b></br></br><br />
<span id="little">We cannot see expression level change of fluorescent protein in SDS-PAGE.</span><br />
</br><br />
<br />
<br />
</div><br />
<br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="31"><br />
<div class="date">August 31<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
</br></br><br />
</div><br />
</div><br />
<br />
</body><br />
</html><br />
{{:Team:KAIST_Korea/footer}}<br />
<br />
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<span id="little">CONTENT</span><br />
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<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_08##Fig#" src="#"></img></div><br />
<div style="clear:both;"></div><br />
<br />
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<br />
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<br />
<br />
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<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<br />
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<br />
<div id="content_note" ><br />
<br />
<span id="little">CONTENT2</span><br />
</br></br><br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="2_08##Fig#" src="#"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br><br />
<br />
<span id="little"></span><br />
<br />
</br><br />
</div><br />
--></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_10
Team:KAIST Korea/Notebook Labnote/2012 10
2012-10-27T01:51:46Z
<p>Mandie: </p>
<hr />
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<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
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<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-October</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
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<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">October</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">October 1<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
<section id="2"><br />
<div class="date">October 2<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">bFMO template preparation with new primer</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/a3/121002.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>We have found that our bFMO sequence was wrong. So that we have ordered new primers with right sequence.</br></span><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="3"><br />
<div class="date">October 3<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Visualization of inversion from GFP to RFP</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little"></br>bfmo gene is constructed in the vector(pJ401, Km registance) and transformed into DH5α.</br>50λ cell from stock is inoculated in 5mL LB </br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Visualization of inversion from GFP to RFP</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
<span id="little">We sampled the cell every 3hr. We induced one group of samples and un-induced another group of samples as a control. We took the picture of cell with confocal microscope at absorbance 500nm for GFP expression and 600nm for RFP expression. </br><br />
</span><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/ae/121003.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">We obtained proper data. Un-induced sample doesn’t show measurable RFP expression throughout the culture. However, induced sample shows the increase of RFP expression. Increase of GFP level in induced sample is due to the stability of GFP and time delay before inversion. </span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Insert DNA preparation with OE pcr</div><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/24/121003-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/87/121003-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
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<div class="date">October 5<sup>th</sup> 2012</div></br><br />
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<span id="little">Regional Jamboree at HKUST!!!!</span><br />
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<span id="little">Regional Jamboree at HKUST!!!!</span><br />
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<div class="date">October 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/>Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Cell culture in 96-well plate for Indigo expression time optimization</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little"></br>We set the O.D value to 0.1 and cultured 1.5ml cell in 96 deep well plate. At each time point, we transferred 1ml cell from 96 deep well plate to EP tube and 200λ cell to flat 96 well plate for O.D check.</br><br />
After cell down in EP tube, we discarded the supernatant and resuspended the pellet in 200λ DMSO. We did sonication to the sample and after another cell down, we took the supernatant for TECAN measurement. Also, we checked absorbance after 3 days incubation and before 3days incubation.<br />
</br>We used TECAN at absorbance 620nm to measure the quantity of indigo production</br></br></span><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/3/3f/Tecan-1.PNG"></img></div><br />
<div style="clear:both;"></div></br><br />
<br />
<div align="center"><img id="figure" src="https://static.igem.org/mediawiki/2012/7/73/Tecan-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<span id="little"></br></br>We subtracted absorbance value of pure LB from value of samples and we set all samples to have same amount of cell using O.D value </br><br />
Graph shown below is indigo intensity per cell. After 3 days incubation, indigo seems to be degraded and all the samples seem to have almost same indigo level.</br></br></span><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/f/fc/Tecan_indigo-1.PNG"></img></div><br />
<div style="clear:both;"></div></br><br />
<br />
<span id="little"></br>We excluded 0hr data from the graph to see teh rising tendency clearly.</br></br></span><br />
<br />
<div align="center"><img id="figure" src="https://static.igem.org/mediawiki/2012/0/06/Tecan_indigo-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<b></br>Discussion</b></br><br />
<br />
<span id="little"></br>This data shows that the cell successfully produces indigo. We can use this gene as a reporter of our module. However, we only checked the increase of indigo. It may decrease after prolonged culture. We should do additional experiments to get the peak in the graph and optimize the production time of indigo. </br></span><br />
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<div class="date">October 19<sup>th</sup> 2012</div></br><br />
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Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_10
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<br />
<div id="menucontent"><br />
<img src="https://static.igem.org/mediawiki/2012/4/4d/Note_ico.png"></img><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Overview"><img src="https://static.igem.org/mediawiki/2012/e/e3/Overview_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Protocol"><img src="https://static.igem.org/mediawiki/2012/b/b6/Protocol_ico.png"/></a><br />
<a href="https://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e0/Labnote_ico.png"/></a><br />
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<br />
<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-October</span></div><br />
<img id="horizontal-line" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img><br />
<br />
<div id="kaistcontent"><br />
<div><br />
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<h1>Labnote</h1><br />
<span id="sub-title">October</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">October 1<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
<section id="2"><br />
<div class="date">October 2<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">bFMO template preparation with new primer</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/a3/121002.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>We have found that our bFMO sequence was wrong. So that we have ordered new primers with right sequence.</br></span><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="3"><br />
<div class="date">October 3<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Visualization of inversion from GFP to RFP</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little"></br>bfmo gene is constructed in the vector(pJ401, Km registance) and transformed into DH5α.</br>50λ cell from stock is inoculated in 5mL LB </br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Visualization of inversion from GFP to RFP</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
<span id="little">We sampled the cell every 3hr. We induced one group of samples and un-induced another group of samples as a control. We took the picture of cell with confocal microscope at absorbance 500nm for GFP expression and 600nm for RFP expression. </br><br />
</span><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/ae/121003.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">We obtained proper data. Un-induced sample doesn’t show measurable RFP expression throughout the culture. However, induced sample shows the increase of RFP expression. Increase of GFP level in induced sample is due to the stability of GFP and time delay before inversion. </span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Insert DNA preparation with OE pcr</div><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/24/121003-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/87/121003-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8a/121003-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="4"><br />
<div class="date">October 4<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="5"><br />
<div class="date">October 5<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">Regional Jamboree at HKUST!!!!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="6"><br />
<div class="date">October 6<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">Regional Jamboree at HKUST!!!!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="7"><br />
<div class="date">October 7<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">Regional Jamboree at HKUST!!!!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="8"><br />
<div class="date">October 8<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="9"><br />
<div class="date">October 9<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="10"><br />
<div class="date">October 10<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="11"><br />
<div class="date">October 11<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="12"><br />
<div class="date">October 12<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
<br />
</section><br />
<section id="13"><br />
<div class="date">October 13<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="14"><br />
<div class="date">October 14<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="15"><br />
<div class="date">October 15<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="16"><br />
<div class="date">October 16<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="17"><br />
<div class="date">October 17<sup>th</sup> 2012</div></br><br />
<br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="18"><br />
<div class="date">October 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Cell culture in 96-well plate for Indigo expression time optimization</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little"></br>We set the O.D value to 0.1 and cultured 1.5ml cell in 96 deep well plate. At each time point, we transferred 1ml cell from 96 deep well plate to EP tube and 200λ cell to flat 96 well plate for O.D check.</br><br />
After cell down in EP tube, we discarded the supernatant and resuspended the pellet in 200λ DMSO. We did sonication to the sample and after another cell down, we took the supernatant for TECAN measurement. Also, we checked absorbance after 3 days incubation and before 3days incubation.<br />
</br>We used TECAN at absorbance 620nm to measure the quantity of indigo production</br></br></span><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/3/3f/Tecan-1.PNG"></img></div><br />
<div style="clear:both;"></div></br><br />
<br />
<div align="center"><img id="figure" src="https://static.igem.org/mediawiki/2012/7/73/Tecan-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<span id="little"></br>We subtracted absorbance value of pure LB from value of samples and we set all samples to have same amount of cell using O.D value </br></br></span><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little"></br>This data shows that the cell successfully produces indigo. We can use this gene as a reporter of our module. However, we only checked the increase of indigo. It may decrease after prolonged culture. We should do additional experiments to get the peak in the graph and optimize the production time of indigo. </br></span><br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
<br />
</section><br />
<section id="19"><br />
<div class="date">October 19<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<br />
<br />
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<br />
</section><br />
<section id="20"><br />
<div class="date">October 20<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
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<br />
</section><br />
<section id="21"><br />
<div class="date">October 21<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
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</section><br />
<section id="22"><br />
<div class="date">October 22<sup>nd</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
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</section><br />
<section id="23"><br />
<div class="date">October 23<sup>rd</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
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<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="24"><br />
<div class="date">October 24<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
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</section><br />
<section id="25"><br />
<div class="date">October 25<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="26"><br />
<div class="date">October 26<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="27"><br />
<div class="date">October 27<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="28"><br />
<div class="date">October 28<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="29"><br />
<div class="date">October 29<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
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</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="30"><br />
<div class="date">October 30<sup>th</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
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</section><br />
<section id="31"><br />
<div class="date">October 31<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
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<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
</br></br><br />
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{{:Team:KAIST_Korea/footer}}</div>
Mandie
http://2012.igem.org/File:Tecan_indigo-2.PNG
File:Tecan indigo-2.PNG
2012-10-27T01:34:26Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/File:Tecan_indigo-1.PNG
File:Tecan indigo-1.PNG
2012-10-27T01:33:56Z
<p>Mandie: </p>
<hr />
<div></div>
Mandie
http://2012.igem.org/Team:KAIST_Korea/Notebook_Labnote/2012_10
Team:KAIST Korea/Notebook Labnote/2012 10
2012-10-27T01:25:26Z
<p>Mandie: </p>
<hr />
<div>{{:Team:KAIST_Korea/top_home}}<br />
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<div id="top-img-description-box"><span style="font-size:22px;" id="top-img-description">Notebook : Labnote-October</span></div><br />
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<br />
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<img id="starter-grad" style="height:80px;" src="https://static.igem.org/mediawiki/2012/9/95/Starter_gradient_kaist.png"></img><br />
<h1>Labnote</h1><br />
<span id="sub-title">October</span></br></br><br />
</div><br />
<br />
<section id="1"><br />
<div class="date">October 1<sup>st</sup> 2012</div></br><br />
<div id="content_note" ><br />
<span id="little">No Special Event!</span><br />
</br><br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<br />
<section id="2"><br />
<div class="date">October 2<sup>nd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">bFMO template preparation with new primer</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/a3/121002.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<span id="little"></br>We have found that our bFMO sequence was wrong. So that we have ordered new primers with right sequence.</br></span><br />
<br />
<br />
<br />
</div><br />
<a id="back_to_the_calender" style="font-size:12px;color:#111;text-decoration:none;float:right;margin:20px 0px 0px 5px;" href="https://2012.igem.org/wiki/index.php?title=Team:KAIST_Korea/Notebook_Labnote"><img src="https://static.igem.org/mediawiki/2012/e/e3/KAIST_calendar_icon.png" style="width:60px;height:60px;display:none;"/>Back to the Calendar</a><br />
</section><br />
<section id="3"><br />
<div class="date">October 3<sup>rd</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Visualization of inversion from GFP to RFP</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little"></br>bfmo gene is constructed in the vector(pJ401, Km registance) and transformed into DH5α.</br>50λ cell from stock is inoculated in 5mL LB </br></span><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Visualization of inversion from GFP to RFP</div><br />
<br />
<div id="content_note" ><br />
<b>Procedure</b></br></br><br />
<span id="little">We sampled the cell every 3hr. We induced one group of samples and un-induced another group of samples as a control. We took the picture of cell with confocal microscope at absorbance 500nm for GFP expression and 600nm for RFP expression. </br><br />
</span><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/a/ae/121003.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</br></br><br />
<b>Discussion</b></br></br><br />
<span id="little">We obtained proper data. Un-induced sample doesn’t show measurable RFP expression throughout the culture. However, induced sample shows the increase of RFP expression. Increase of GFP level in induced sample is due to the stability of GFP and time delay before inversion. </span><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Insert DNA preparation with OE pcr</div><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/2/24/121003-1.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
</div><br />
<br />
</br><div align="center"><img style="width:670px; height:2px;" src="https://static.igem.org/mediawiki/2012/7/7f/Horizontal_line_kaist.png"></img></div></br><br />
<div class="note-title"> Cloning</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/87/121003-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/8/8a/121003-3.PNG"></img></div><br />
<div style="clear:both;"></div><br />
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</section><br />
<section id="4"><br />
<div class="date">October 4<sup>th</sup> 2012</div></br><br />
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<section id="5"><br />
<div class="date">October 5<sup>th</sup> 2012</div></br><br />
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<div class="date">October 6<sup>th</sup> 2012</div></br><br />
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<section id="7"><br />
<div class="date">October 7<sup>th</sup> 2012</div></br><br />
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<div class="date">October 8<sup>th</sup> 2012</div></br><br />
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<div class="date">October 9<sup>th</sup> 2012</div></br><br />
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<div class="date">October 10<sup>th</sup> 2012</div></br><br />
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<div class="date">October 11<sup>th</sup> 2012</div></br><br />
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<div class="date">October 15<sup>th</sup> 2012</div></br><br />
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<div class="date">October 17<sup>th</sup> 2012</div></br><br />
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</section><br />
<section id="18"><br />
<div class="date">October 18<sup>th</sup> 2012</div></br><br />
<div id="linegray" align="center" style="font-size:15px; font-weight:bold; color:#888;"><img style="height:3px;width:269px;margin:0px 5px 0px 0px; "src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/><img style="width:40px; height:28px;float:bottom;"id="flipflop" src='https://static.igem.org/mediawiki/2012/2/22/KAIST_FlipFlop.png'/> Flip Flop<img style="height:3px;width:269px; margin:0px 0px 0px 5px;"src="https://static.igem.org/mediawiki/2012/1/1b/KAIST_horizontal_gray.png"/></div></br><br />
<div class="note-title">Cell culture in 96-well plate for Indigo expression time optimization</div><br />
<br />
<div id="content_note" ><br />
<br />
<b>Procedure</b></br></br><br />
<br />
<span id="little"></br>We set the O.D value to 0.1 and cultured 1.5ml cell in 96 deep well plate. At each time point, we transferred 1ml cell from 96 deep well plate to EP tube and 200λ cell to flat 96 well plate for O.D check.</br><br />
After cell down in EP tube, we discarded the supernatant and resuspended the pellet in 200λ DMSO. We did sonication to the sample and after another cell down, we took the supernatant for TECAN measurement. Also, we checked absorbance after 3 days incubation and before 3days incubation.<br />
</br>We used TECAN at absorbance 620nm to measure the quantity of indigo production</br></br></span><br />
<br />
<b>Results</b></br></br><br />
<br />
<div align="center"><img id="figure" alt="0818Fig1" src="https://static.igem.org/mediawiki/2012/3/3f/Tecan-1.PNG"></img></div><br />
<div style="clear:both;"></div></br><br />
<br />
<div align="center"><img id="figure" src="https://static.igem.org/mediawiki/2012/7/73/Tecan-2.PNG"></img></div><br />
<div style="clear:both;"></div><br />
<br />
<br />
<span id="little"></br>We subtracted absorbance value of pure LB from value of samples and we set all samples to have same amount of cell using O.D value </br></br></span><br />
<br />
<b>Discussion</b></br></br><br />
<br />
<span id="little"></br>This data shows that the cell successfully produces indigo. We can use this gene as a reporter of our module. However, we only checked the increase of indigo. It may decrease after prolonged culture. We should do additional experiments to get the peak in the graph and optimize the production time of indigo. </br></span><br />
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<section id="19"><br />
<div class="date">October 19<sup>th</sup> 2012</div></br><br />
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<div class="date">October 20<sup>th</sup> 2012</div></br><br />
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<div class="date">October 21<sup>st</sup> 2012</div></br><br />
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<div class="date">October 22<sup>nd</sup> 2012</div></br><br />
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<div class="date">October 23<sup>rd</sup> 2012</div></br><br />
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<div class="date">October 24<sup>th</sup> 2012</div></br><br />
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<div class="date">October 25<sup>th</sup> 2012</div></br><br />
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<div class="date">October 29<sup>th</sup> 2012</div></br><br />
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<div class="date">October 30<sup>th</sup> 2012</div></br><br />
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<div class="date">October 31<sup>st</sup> 2012</div></br><br />
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