Team:Grenoble/Biology/Notebook/July/week 30

From 2012.igem.org

(Difference between revisions)
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<body id="Biology">
<body id="Biology">
 +
<div id="cadre">
<div id="cadre">
<section>
<section>
-
<h1>July</h1>
+
<h1>August</h1>
-
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/July">Week 27</a> •  
+
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/August">Week 31</a> •
-
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/July/week_28">Week 28</a> •  
+
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/July/week_32">Week 32</a> •  
-
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/July/week_29">Week 29</a> •  
+
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/July/week_33">Week 33</a> •  
-
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/July/week_30">Week 30</a>
+
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/July/week_34">Week 34</a> •  
 +
<a href="https://2012.igem.org/Team:Grenoble/Biology/Notebook/July/week_35">Week 35</a>
</section>
</section>
<br/>
<br/>
<br/>
<br/>
<section>
<section>
-
<h1> Week 30: July 23<span class="exposant">rd</span> to 29<span class="exposant">th</span> </h1>
+
<h1> Week 32: August 06<span class="exposant">th</span> to 12<span class="exposant">th</span> </h1>
<h2> Goal of the week: </h2>
<h2> Goal of the week: </h2>
-
We wanted to test the Gibson Assembly and the transformation protocols, recover and amplify some biobricks involved in our genetic networks:
 
-
<ul>
 
-
<li>pAra/Bad_RBS_GFP (1300bp)</li>
 
-
<li>RBS_Cya (2600bp)</li>
 
-
<li>fha (80bp)</li>
 
-
<li>eCFP (800bp)</li>
 
-
<li>pSB4K5 (2400bp)</li>
 
-
</ul>
 
-
<br/>
 
-
We also wanted to test the "AND" gate (CRP°cAMP - araC) without the amplifier module.<br/>
 
</section>
</section>
<br/>
<br/>
<section>
<section>
-
<h2> Monday, July 23<span class="exposant">rd</span>:</h2>
+
<h2> Monday, August 06<span class="exposant">th</span>:</h2>
-
We did an experiment (protocol) in order to test the "AND" gate (CRP°cAMP - araC) without the amplifier module. <br/>
+
We wanted to check if the Gibson Assemblies (12/08/01) worked well.<br/>
<br/>
<br/>
-
<center><img src="https://static.igem.org/mediawiki/2012/2/2d/AND-gate.png" alt="results_AND_Gate"/></center>
+
To separate the GA miniprep (12/08/03) products, we prepared a 1.8% TAE agarose gel.<br/>
-
<div class="legend">strain used = BW25113 cya<span class="exposant">-</span> transformed with pAra/Bad_RBS_GFP<br/></div>
+
-
<br/>The experiments worked well, the "AND" gate worked as expected.<br/>
+
-
<br/>
+
-
We did PCRs with HF Phusion enzyme (protocol) on miniprep in order to amplify pAra/Bad_RBS_GFP, and on colony to amplify RBS_Cya. We did the PCR both with and without DMSO.<br/>
+
-
<br/>
+
-
To separate the PCR products (12/07/23 and 12/07/20), we prepared a 1.8% TAE agarose gel.<br/>
+
Migration conditions = 100V during 30 min.<br/>
Migration conditions = 100V during 30 min.<br/>
In order to reveal the DNA fragments, we used EtBr.<br/>
In order to reveal the DNA fragments, we used EtBr.<br/>
<br/>
<br/>
-
<center><img src="https://static.igem.org/mediawiki/2012/c/c1/120723.jpg" alt="photo_gel_16"/></center>
+
<center><img src="https://static.igem.org/mediawiki/2012/4/44/120806_pcr_miniprep_2.jpg" alt="photo_gel_27"/></center>
-
<div class="legend"><u>Migration result for a 1.3% TAE agarose gel</u><br/>
+
-
  <i>(the DNA ladder scale is in kb)</i><br/>
+
-
Lane 1: DNA ladder 80pb-10kb (fermentas)<br/>
+
-
Lane 2: pSB4K5 PCR product (DMSO)<br/>
+
-
Lane 3: fha1 PCR product (DMSO)<br/>
+
-
Lane 4: fha1 PCR product<br/>
+
-
Lane 5: pSB4K5 PCR product<br/>
+
-
Lane 6: pAra/Bad_RBS_GFP PCR product (DMSO)<br/>
+
-
Lane 7: pAra/Bad_RBS_GFP PCR product (DMSO)<br/>
+
-
Lane 8: RBS_Cya PCR product (DMSO)<br/>
+
-
Lane 9: RBS_Cya PCR product<br/>
+
-
Lane 10: pAra/Bad_RBS_GFP PCR product<br/>
+
-
Lane 11: pAra/Bad_RBS_GFP PCR product<br/>
+
-
Lane 12: DNA ladder 80bp-10kb (fermentas)<br/></div>
+
-
<br/>
+
-
We saw primer dimer bands and the bands corresponding to the amplified fha1 sequence (lanes 3 & 4). For the other, there was a PCR condition problem.<br/>
+
-
<br/>
+
-
We realised a DNA extraction (protocol kit: Nucleospin extract II) from the two fha PCR products <span class="code">120723AM_PCR_020</span>.<br/>
+
-
<br/>
+
-
We did touchdown PCRs with HF Phusion enzyme (protocol) on digestion product (eCFP E0022 and E0422) (12/07/20) in order to amplify eCFP. We did the PCR both with and without DMSO.<br/>
+
-
<br/>
+
-
Using iGEM 2012 biobricks we transformed (new protocol) BW25113 WT competent cells (protocol) with pSB4K5.<br/>
+
-
</section>
+
-
<br/>
+
-
<section>
+
-
<h2> Tuesday, July 24<span class="exposant">th</span>:</h2>
+
-
To separate the PCR products (12/07/23), we prepared a 1.3% TAE agarose gel.<br/>
+
-
Migration conditions = 100V during 30 min.<br/>
+
-
In order to reveal the DNA fragments, we used EtBr.<br/>
+
-
<br/>
+
-
<center><img src="https://static.igem.org/mediawiki/2012/e/e0/120724_%282%29.jpg" alt="photo_gel_17"/></center>
+
-
<div class="legend"><u>Migration result for a 1.3% TAE agarose gel</u><br/>
+
-
  <i>(the DNA ladder scale is in kb)</i><br/>
+
-
Lane 1: DNA ladder 80bp-10kb (fermentas)<br/>
+
-
Lane 2: E0022 PCR product<br/>
+
-
Lane 3: E0022 PCR product (DMSO)<br/>
+
-
Lane 4: E0422 PCR product<br/>
+
-
Lane 5: E0422 PCR product (DMSO)<br/>
+
-
Lane 6: DNA ladder 80bp-10kb (fermentas)<br/></div>
+
-
<br/>
+
-
We only saw primer dimer bands, there was a PCR condition problem.<br/>
+
-
<br/>
+
-
The transformation with pSB4K5 (12/07/23) showed result (growth on plates).<br/>
+
-
<br/>
+
-
We did some digestions (protocol) on miniprep (12/07/19) to recover eCFP (E0022). The digestions were achieved with two restriction enzymes : XbaI and SpeI during 10 minutes.<br/>
+
-
<br/>
+
-
To separate the digestion products, we prepared a 1.3% TAE agarose gel.<br/>
+
-
Migration conditions = 100V during 30 min.<br/>
+
-
In order to reveal the DNA fragments, we used EtBr.<br/>
+
-
<br/>
+
-
<center><img src="https://static.igem.org/mediawiki/2012/6/6b/120724.jpg" alt="photo_gel_18"/></center>
+
-
<div class="legend"><u>Migration result for a 1.3% TAE agarose gel</u><br/>
+
-
  <i>(the DNA ladder scale is in kb)</i><br/>
+
-
Lane 1: DNA ladder 1kb (biolabs)<br/>
+
-
Lane 2: E0022 digestion product<br/>
+
-
Lane 3: pSB4K5 digestion product<br/>
+
-
Lane 4: DNA ladder 80bp-1kb (fermentas)<br/></div>
+
-
<br/>
+
-
The digestion worked, we saw DNA bands at the expected positions.<br/>
+
-
<br/>
+
-
We did  PCR with HF Phusion enzyme (protocol) from one of these digestion product (eCFP E0022) in order to amplify eCFP and a colony PCR to amplify pSB4K5.<br/>
+
-
<br/>
+
-
To separate the PCR products, we prepared a 1.3% TAE agarose gel.<br/>
+
-
Migration conditions = 100V during 30 min.<br/>
+
-
In order to reveal the DNA fragments, we used EtBr.<br/>
+
-
<br/>
+
-
There was no result (data not shown).<br/>
+
-
<br/>
+
-
We didn’t achieve to amplify pSB4K5. We decided to change it, we chose pSB4C5 instead. So we had to change pSB3C5, we chose pSB3T5. <br/>
+
-
<br/>
+
-
With iGEM 2012 biobricks we transformed (new protocol) BW25113 WT competent cells (protocol) with pSB4C5.<br/>
+
-
<br/>
+
-
We did an overlapping PCR with HF Phusion enzyme (protocol) on the digestion product (eCFP E0022) in order to amplify eCFP and on miniprep to amplify fha1.<br/>
+
-
</section>
+
-
<br/>
+
-
<section>
+
-
<h2> Wednesday, July 25<span class="exposant">th</span>:</h2>
+
-
To separate the PCR products (12/07/24), we prepared a 1.8% TAE agarose gel.<br/>
+
-
Migration conditions = 100V during 30 min.<br/>
+
-
In order to reveal the DNA fragments, we used EtBr.<br/>
+
-
<br/>
+
-
There was a migration problem (data not shown).<br/>
+
-
<br/>
+
-
The transformation with pSB4C5 (12/07/24) was successful (growth on plates). We decided to relaunch it in fresh liquid LB.<br/>
+
-
<br/>
+
-
We did some glycerol stocks :
+
-
<ul>
+
-
<li>GA: pLAC_RBS_RsmA <span class="code">120725GH_001</span></li>
+
-
<li>pSB4K5 <span class="code">120725GH_002</span></li>
+
-
<li>E0422 (eCFP) <span class="code">120725GH_003</span></li>
+
-
<li>LuxI <span class="code">120725GH_004</span></li>
+
-
<li>E0022 (eCFP) <span class="code">120725GH_005</span></li>
+
-
<li>LuxR <span class="code">120725GH_006</span></li>
+
-
<li>pLux <span class="code">120725GH_007</span></li>
+
-
<li>I13601 <span class="code">120725GH_008</span></li>
+
-
</ul>
+
-
<br/>
+
-
We did a colony PCR with GoTaq enzyme (protocol) in order to amplify pSB4C5.<br/>
+
-
<br/>
+
-
To separate the PCR products (07/24), we prepared a 1.8% TAE agarose gel.<br/>
+
-
Migration conditions = 100V during 30 min.<br/>
+
-
In order to reveal the DNA fragments, we used EtBr.<br/>
+
-
<br/>
+
-
<center><img src="https://static.igem.org/mediawiki/2012/3/3a/120725_%28PCR_colonie_pSB4C5%29.jpg" alt="photo_gel_19"/></center>
+
<div class="legend"><u>Migration result for a 1.8% TAE agarose gel</u><br/>
<div class="legend"><u>Migration result for a 1.8% TAE agarose gel</u><br/>
   <i>(the DNA ladder scale is in kb)</i><br/>
   <i>(the DNA ladder scale is in kb)</i><br/>
-
Lane 1: DNA ladder 1kb (biolabs)<br/>
+
<ul text-align:"left">Lane 1: pLAC_rsmY (pSB1A3) <b>3</b> miniprep product<br/>
-
Lane 2: pSB4C5 PCR product<br/>
+
Lane 2: pLAC_rsmY (pSB1A3) <b>2</b> miniprep product<br/>
-
Lane 3: DNA ladder 1kb (biolabs)<br/></div>
+
Lane 3: pLAC_rsmY (pSB1A3) <b>1</b> miniprep product<br/>
 +
Lane 4: pLAC_fha1_eCFP (pSB4C5) <b>2</b> miniprep product<br/>
 +
Lane 5: pLAC_fha1_eCFP (pSB4C5) <b>1</b> miniprep product<br/>
 +
Lane 6: DNA ladder 1kb (biolabs)<br/>
 +
Lane 7: pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) <b>1</b> miniprep product<br/>
 +
Lane 8: pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) <b>2</b> miniprep product<br/>
 +
Lane 9: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>4</b> miniprep product<br/>
 +
Lane 10: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>3</b> miniprep product<br/>
 +
Lane 11: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>1</b> miniprep product<br/>
 +
Lane 12: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>2</b> miniprep product<br/>
 +
Lane 13: DNA ladder 1kb (biolabs)<br/></ul></div>
<br/>
<br/>
-
We concluded that the pSB4C5 amplification worked well, so we decided to do the PCR again in order to do a DNA extraction.<br/>
 
-
</section>
 
-
<br/>
 
-
<section>
 
-
<h2> Thursday, July 26<span class="exposant">th</span>:</h2>
 
-
We did glycerol stock of the strain transformed with pSB4C5 <span class="code">120726AM_009</span>.<br/>
 
-
<br/>
 
-
We did a colony PCR withs HF Phusion Enzyme (protocol) in order to amplify pSB4C5.<br/>
 
-
<br/>
 
-
We did a miniprep (protocol kit: Nucleospin plasmid Quick Pure) on the transformed strain with pSB4K5, on which we wanted to recover pSB4K5.<br/>
 
<br/>
<br/>
-
We did some digestions (protocol) on this miniprep, in order to check if pSB4K5 was ok. The digestion was realised with two restriction enzymes : XbaI and PstI during 10 minutes.<br/>
+
We did some digestions (protocol) on GA miniprep (12/08/03). The digestions were achieved with XbaI during 10 minutes.<br/>
<br/>
<br/>
-
To separate the PCR products (12/07/24), we prepared a 1.8% TAE agarose gel.<br/>
+
To separate the digestion products, we prepared two 1.8% TAE agarose gel.<br/>
Migration conditions = 100V during 30 min.<br/>
Migration conditions = 100V during 30 min.<br/>
In order to reveal the DNA fragments, we used EtBr.<br/>
In order to reveal the DNA fragments, we used EtBr.<br/>
<br/>
<br/>
-
<center><img src="https://static.igem.org/mediawiki/2012/6/66/120726.jpg" alt="photo_gel_20"/></center>
+
<center><img src="https://static.igem.org/mediawiki/2012/c/cd/120806_dig2.jpg" alt="photo_gel_28"/></center>
<div class="legend"><u>Migration result for a 1.8% TAE agarose gel</u><br/>
<div class="legend"><u>Migration result for a 1.8% TAE agarose gel</u><br/>
   <i>(the DNA ladder scale is in kb)</i><br/>
   <i>(the DNA ladder scale is in kb)</i><br/>
-
Lane 1: DNA ladder 1kb (biolabs)<br/>
+
Lane 1: data not shown pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) <b>1</b> digestion product<br/>
-
Lanes 2 and 3: pSB4K5 without digestion<br/>
+
Lane 2: data not shown pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) <b>1</b> miniprep product<br/>
-
Lanes 4 and 5: pSB4K5 digestion product <br/>
+
Lane 3: pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) <b>2</b> digestion product<br/>
-
Lane 6: empty lane<br/>
+
Lane 4: pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) <b>2</b> miniprep product<br/>
-
Lanes 7 and 8: pSB4C5 PCR product<br/>
+
Lane 5: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>4</b> digestion product<br/>
-
Lane 9: DNA ladder 1kb (biolabs)<br/></div>
+
Lane 6: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>4</b> miniprep product<br/>
 +
Lane 7: DNA ladder 1kb (biolabs)<br/>
 +
Lane 8: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>3</b> digestion product<br/>
 +
Lane 9: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>3</b> miniprep product<br/>
 +
Lane 10: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>1</b> digestion product<br/>
 +
Lane 11: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>1</b> miniprep product<br/>
 +
Lane 12: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>2</b> digestion product<br/>
 +
Lane 13: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>2</b> miniprep product<br/></div>
<br/>
<br/>
-
We concluded that the digestion worked well (we seen the expected DNA bands), and so was the PCR on pSB4C5.<br/>
+
 
<br/>
<br/>
-
We realised a DNA extraction (protocol kit: Nucleospin extract II) on the two pSB4C5 PCR products <span class="code">120726AM_PCR_021</span>.<br/>
 
<br/>
<br/>
-
We didn’t achieve to amplify eCFP, we decided to change it. We chose mcherry and GFP.<br/>
+
<center><img src="https://static.igem.org/mediawiki/2012/3/3e/120806.jpg" alt="photo_gel_29"/></center>
-
<br/>
+
-
We transformed  (new protocol) BW25113 WT competent cells (protocol) with biobricks from iGEM 2012 kit :
+
-
<ul>
+
-
<li>BBa_I763011 (GFP) </li>
+
-
<li>BBa_J06702 (mcherry) (in duplicate : in LB with Ampicillin and in LB with Ampicillin and Kanamycin)</li>
+
-
<li>pSB1A3</li>
+
-
<li>BBa_R0082 (pompC)</li>
+
-
<li>BBa_J23119 (pConst)</li>
+
-
</ul>
+
-
</section>
+
-
<br/>
+
-
<section>
+
-
<h2> Friday, July 27<span class="exposant">th</span>:</h2>
+
-
The five transformations (12/07/26) were successful (growth on the expected plates). We did glycerol stocks of:
+
-
<ul>
+
-
<li>BBa_J06702 (mcherry in LB with Ampicillin and Kanamycin) <span class="code">120727GH_010</span></li>
+
-
<li>pSB1A3 <span class="code">120727GH_011</span></li>
+
-
<li>BBa_J23119 (pConst) <span class="code">120727GH_012</span></li>
+
-
<li>BBa_R0082 (pOmpC) <span class="code">120727GH_013</span></li>
+
-
<li>BBa_J06702 (mcherry in LB with Ampicillin) <span class="code">120727GH_014</span></li>
+
-
<li>BBa_I763011 (GFP) <span class="code">120727GH_015</span></li>
+
-
</ul>
+
-
<br/>
+
-
We did some colony PCRs with HF Phusion enzyme (protocol) in order to amplify RBS_Cya (on BW25113 WT strain), pAra/Bad_RBS_GFP (on BW25113 Cya- pAra/Bad) and pSB4C5 (on 12/07/25 colony).<br/>
+
-
<br/>
+
-
To separate the PCR products, we prepared a 1.3% TAE agarose gel.<br/>
+
-
Migration conditions = 100V during 30 min.<br/>
+
-
In order to reveal the DNA fragments, we used EtBr.<br/>
+
-
<br/>
+
-
<center><img src="https://static.igem.org/mediawiki/2012/b/b1/120727.jpg" alt="photo_gel_21"/></center>
+
<div class="legend"><u>Migration result for a 1.8% TAE agarose gel</u><br/>
<div class="legend"><u>Migration result for a 1.8% TAE agarose gel</u><br/>
   <i>(the DNA ladder scale is in kb)</i><br/>
   <i>(the DNA ladder scale is in kb)</i><br/>
Lane 1: DNA ladder 1kb (biolabs)<br/>
Lane 1: DNA ladder 1kb (biolabs)<br/>
-
Lanes 2 and 3: pAra/Bad_RBS_GFP PCR product<br/>
+
Lane 2: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>4</b> digestion product<br/>
-
Lanes 4 and 5: RBS_Cya PCR product<br/>
+
Lane 3: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>4</b> miniprep product<br/>
-
Lanes 6 and 7: pSB4C5 PCR product<br/>
+
Lane 4: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>1</b> digestion product<br/>
-
Lane 8: DNA ladder 1kb (biolabs)<br/></div>
+
Lane 5: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>1</b> miniprep product<br/>
-
 
+
Lane 6: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>2</b> digestion product<br/>
-
 
+
Lane 7: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) <b>2</b> miniprep product<br/>
 +
Lane 8:DNA ladder 80pb-10kb (fermentas)<br/>
 +
</div>
<br/>
<br/>
-
The pSB4C5 amplification didn’t work. However, the RBS_Cya and pAra/Bad_RBS_GFP amplications worked.<br/>
 
-
<br/>
 
-
We realised a DNA extraction (protocol kit: Nucleospin extract II) from the two pAra/Bad_RBS_GFP PCR products <span class="code">120727AM_PCR_023</span> and the two Cya PCR products <span class="code">120727AM_PCR_022</span>.<br/>
 
</section>
</section>
<br/>
<br/>
<section>
<section>
-
<h2> Conclusion of the week:</h2>
 
-
We replaced pSB4K5 with pSB4C5 and pSB3C5 with pSB3T5. We also replaced eCFP with mcherry or GFP.<br/>
 
-
<br/>We have achieved to amplify pAra/Bad_RBS_GFP // RBS_Cya // fha // pSB4C5 (fha) and we began the experiments in order to recover pOmpC (for the detection module) and pConst.<br/>
 
-
<br/>The "AND" gate worked as expected.
 
-
</section>
 
-
<br/>
 
-
<br/>
 
</div>
</div>
</body>
</body>

Revision as of 13:12, 20 August 2012

iGEM Grenoble 2012

iGEM Grenoble iGEM
iGEM 2012
main page
Project

August

Week 31 • Week 32 • Week 33 • Week 34 • Week 35


Week 32: August 06th to 12th

Goal of the week:


Monday, August 06th:

We wanted to check if the Gibson Assemblies (12/08/01) worked well.

To separate the GA miniprep (12/08/03) products, we prepared a 1.8% TAE agarose gel.
Migration conditions = 100V during 30 min.
In order to reveal the DNA fragments, we used EtBr.

photo_gel_27
Migration result for a 1.8% TAE agarose gel
(the DNA ladder scale is in kb)
    Lane 1: pLAC_rsmY (pSB1A3) 3 miniprep product
    Lane 2: pLAC_rsmY (pSB1A3) 2 miniprep product
    Lane 3: pLAC_rsmY (pSB1A3) 1 miniprep product
    Lane 4: pLAC_fha1_eCFP (pSB4C5) 2 miniprep product
    Lane 5: pLAC_fha1_eCFP (pSB4C5) 1 miniprep product
    Lane 6: DNA ladder 1kb (biolabs)
    Lane 7: pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) 1 miniprep product
    Lane 8: pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) 2 miniprep product
    Lane 9: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 4 miniprep product
    Lane 10: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 3 miniprep product
    Lane 11: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 1 miniprep product
    Lane 12: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 2 miniprep product
    Lane 13: DNA ladder 1kb (biolabs)


We did some digestions (protocol) on GA miniprep (12/08/03). The digestions were achieved with XbaI during 10 minutes.

To separate the digestion products, we prepared two 1.8% TAE agarose gel.
Migration conditions = 100V during 30 min.
In order to reveal the DNA fragments, we used EtBr.

photo_gel_28
Migration result for a 1.8% TAE agarose gel
(the DNA ladder scale is in kb)
Lane 1: data not shown pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) 1 digestion product
Lane 2: data not shown pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) 1 miniprep product
Lane 3: pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) 2 digestion product
Lane 4: pAra/Bad_RBS_GFP_RBS_Cya (pSB3C5) 2 miniprep product
Lane 5: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 4 digestion product
Lane 6: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 4 miniprep product
Lane 7: DNA ladder 1kb (biolabs)
Lane 8: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 3 digestion product
Lane 9: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 3 miniprep product
Lane 10: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 1 digestion product
Lane 11: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 1 miniprep product
Lane 12: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 2 digestion product
Lane 13: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 2 miniprep product



photo_gel_29
Migration result for a 1.8% TAE agarose gel
(the DNA ladder scale is in kb)
Lane 1: DNA ladder 1kb (biolabs)
Lane 2: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 4 digestion product
Lane 3: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 4 miniprep product
Lane 4: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 1 digestion product
Lane 5: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 1 miniprep product
Lane 6: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 2 digestion product
Lane 7: pAra/Bad_RBS_GFP_RBS_Cya (pSB4C5) 2 miniprep product
Lane 8:DNA ladder 80pb-10kb (fermentas)


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