Team:KIT-Kyoto/Notebook-week3p
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+ | <table border="0" width="965px" align="center"><tr><td width="165px" valign="top" | ||
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+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week1p"><img src="https://static.igem.org/mediawiki/2012/1/1b/Side_week1kit.jpg" width="150" height="30"></a></li> | ||
+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week2p"><img src="https://static.igem.org/mediawiki/2012/9/94/Side_week2kit.jpg" width="150" height="30"></a></li> | ||
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+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week4p"><img src="https://static.igem.org/mediawiki/2012/3/31/Side_week4kit.jpg" width="150" height="30"></a></li> | ||
+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week5p"><img src="https://static.igem.org/mediawiki/2012/e/e7/Side_week5kit.jpg" width="150" height="30"></a></li> | ||
+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week6p"><img src="https://static.igem.org/mediawiki/2012/e/e8/Side_week6kit.jpg" width="150" height="30"></a></li> | ||
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+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week2"><img src="https://static.igem.org/mediawiki/2012/9/94/Side_week2kit.jpg" width="150" height="30"></a></li> | ||
+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week3"><img src="https://static.igem.org/mediawiki/2012/e/e0/Side_week3kit.jpg" width="150" height="30"></a></li> | ||
+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week4"><img src="https://static.igem.org/mediawiki/2012/3/31/Side_week4kit.jpg" width="150" height="30"></a></li> | ||
+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week5"><img src="https://static.igem.org/mediawiki/2012/e/e7/Side_week5kit.jpg" width="150" height="30"></a></li> | ||
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+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week7"><img src="https://static.igem.org/mediawiki/2012/b/b2/Side_weekkit7.jpg" width="150" height="30"></a></li> | ||
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+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-Meeting-june"><img src="https://static.igem.org/mediawiki/2012/9/92/Side_junekit.jpg" width="150" height="30"></a></li> | ||
+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-Meeting-july"><img src="https://static.igem.org/mediawiki/2012/f/f2/Side_julykit.jpg" width="150" height="30"></a></li> | ||
+ | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-Meeting-august"><img src="https://static.igem.org/mediawiki/2012/f/fa/Side_augkit.jpg" width="150" height="30"></a></li><li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-Meeting-september"><img src="https://static.igem.org/mediawiki/2012/8/8b/Side_sepkit.jpg" width="150" height="30"></a></li> | ||
+ | </ul> | ||
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+ | </li> | ||
+ | </ul> | ||
+ | </div> | ||
+ | <br> | ||
+ | </td> | ||
+ | |||
+ | <td width="800px" height="510px" valign="top"> | ||
+ | <div id="MIGI"> | ||
+ | <h2>September 6th</h2> | ||
+ | <br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> All samples </Td></Tr> | ||
+ | <Tr><Td> DNA template </Td><Td> 0.25uL </Td></Tr> | ||
+ | <tr><td> 10× KOD plus buffer </td><td> 5uL </td></tr> | ||
+ | <Tr><Td> 2mM dNTPs </Td><Td> 5uL </Td></Tr> | ||
+ | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr><tr> | ||
+ | <td> 10P 5’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> KOD plus </td><td> 1uL </td></tr> | ||
+ | <Tr><td> dH<sub>2</sub>O </td><td> 34.15uL </td></tr> | ||
+ | <Tr><td> Total </td><td> 50uL </td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> | ||
+ | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> | ||
+ | <Tr><Td> 95℃ </Td><Td> 15sec</Td><Td> 30 cycle </Td></Tr> | ||
+ | <tr><td> 58℃ </td><td> 30sec</td><Td> 30 cycle </Td></tr><tr> | ||
+ | <td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td> 30 cycle </Td></tr> | ||
+ | <tr><td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td></Td></tr> | ||
+ | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/5/53/0906kit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> pSB1C3(XbaⅠ and SpeⅠ cut) </Td><Td> 1uL </Td></Tr> | ||
+ | <Tr><Td> GAL4(XbaⅠ and SpeⅠ cut) </Td><Td> 1uL </Td></Tr> | ||
+ | <Tr><td> dH<sub>2</sub>O </td><td> 3uL </td></tr> | ||
+ | <Tr><Td> Ligation high </Td><Td> 2.5uL </Td></Tr> | ||
+ | <Tr><td> Total </td><td> 7.5uL </td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <h2>September 7th</h2> | ||
+ | <br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> All samples </Td></Tr> | ||
+ | <Tr><Td> DNA template </Td><Td> 0.25uL </Td></Tr> | ||
+ | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td></Tr> | ||
+ | <Tr><td> 2mM dNTPs </td><td> 5uL </td></tr> | ||
+ | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr> | ||
+ | <Tr><td> 10P 5’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> KOD plus </td><td> 1uL </td></tr> | ||
+ | <Tr><td> dH<sub>2</sub>O </td><td> 34.15uL </td></tr> | ||
+ | <Tr><td> Total </td><td> 50uL </td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> | ||
+ | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> | ||
+ | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> | ||
+ | <tr><td> 58℃ </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> | ||
+ | <td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td> 30 cycle </Td></tr> | ||
+ | <tr><td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td></Td></tr> | ||
+ | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/4/4c/0907akit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | <img src="https://static.igem.org/mediawiki/2012/6/68/0907bkit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | <img src="https://static.igem.org/mediawiki/2012/9/9e/0907ckit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <h2>September 8th</h2> | ||
+ | <br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> All samples </Td></Tr> | ||
+ | <Tr><Td> DNA template </Td><Td> 1uL </Td></Tr> | ||
+ | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td></Tr> | ||
+ | <Tr><td> 2mM dNTPs </td><td> 5uL </td></tr> | ||
+ | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr> | ||
+ | <Tr><td> 10P 5’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> KOD plus </td><td> 1uL </td></tr> | ||
+ | <Tr><td> dH<sub>2</sub>O </td><td> 33.4uL </td></tr> | ||
+ | <Tr><td> Total </td><td> 50uL </td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> | ||
+ | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> | ||
+ | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> | ||
+ | <tr><td> 58℃ </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> | ||
+ | <td> 68℃ </td></td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td> 30 cycle </Td></tr> | ||
+ | <tr><td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td></Td></tr> | ||
+ | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/b/be/0908kit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> All samples </Td></Tr> | ||
+ | <Tr><Td> DNA template </Td><Td> 1uL </Td></Tr> | ||
+ | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td></Tr> | ||
+ | <Tr><td> 2mM dNTPs </td><td> 5uL </td></tr> | ||
+ | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr> | ||
+ | <Tr><td> 10P 5’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> KOD plus </td><td> 1uL </td></tr> | ||
+ | <Tr><td> dH<sub>2</sub>O </td><td> 33.4uL </td></tr> | ||
+ | <Tr><td> Total </td><td> 50uL </td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> | ||
+ | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> | ||
+ | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> | ||
+ | <tr><td> 55℃(-1) or 58℃(-2) </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> | ||
+ | <td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td> 30 cycle </Td></tr> | ||
+ | <tr><td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td></Td></tr> | ||
+ | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <h2>September 9th</h2> | ||
+ | <BR> | ||
+ | <img src="https://static.igem.org/mediawiki/2012/1/10/0909akit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> UAS </Td><Td> 40uL </Td></Tr> | ||
+ | <Tr><Td> 4 buffer(NEB) </Td><Td> 5uL </Td></Tr> | ||
+ | <Tr><Td> EcoRⅠ-HF(NEB) </Td><Td> 0.5uL </Td></Tr> | ||
+ | <Tr><td> SpeⅠ(NEB </td><td> 0.5uL </td></tr> | ||
+ | <Tr><Td> 100×BSA </Td><Td> 0.5uL </Td></Tr> | ||
+ | <Tr><td> dH<sub>2</sub>O </td><td> 3.5uL </td></tr> | ||
+ | <Tr><td> Total </td><td> 50uL </td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/9/91/0909bkit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> All sample </Td></Tr> | ||
+ | <Tr><Td> DNA template </Td><Td> 1uL </Td></Tr> | ||
+ | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td></Tr> | ||
+ | <Tr><td> 2mM dNTPs <td> 5uL </td></tr> | ||
+ | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr> | ||
+ | <Tr><td> 10P 5’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> | ||
+ | <Tr><td> KOD plus </td><td> 1uL </td></tr> | ||
+ | <Tr><td> dH<sub>2</sub>O </td><td> 33.4uL </td></tr> | ||
+ | <Tr><td> Total </td><td> 50uL </td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> | ||
+ | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> | ||
+ | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> | ||
+ | <tr><td> 58℃ </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> | ||
+ | <td> 68℃ </td><td> 2min10sec </td><Td> 30 cycle </Td></tr> | ||
+ | <tr><td> 68℃ </td><td> 2min10sec </td><Td></Td></tr> | ||
+ | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/b/bb/0909ckit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> 1uL </Td></Tr> | ||
+ | <Tr><Td></Td><Td> 1uL </Td></Tr> | ||
+ | <Tr><Td> dH<sub>2</sub>O </Td><Td> 3uL </Td></Tr> | ||
+ | <Tr><td> Ligation high </td><td> 2.5uL </td></tr> | ||
+ | <Tr><Td> Total </Td><Td> 7.5uL </Td></Tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> 1uL </Td></Tr> | ||
+ | <Tr><Td> dH<sub>2</sub>O </Td><Td> 4uL </Td></Tr> | ||
+ | <Tr><td> Ligation high </td><td> 2.5uL </td></tr> | ||
+ | <Tr><Td> Total </Td><Td> 7.5uL </Td></Tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <h2>September 10th</h2> | ||
+ | <br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> HS or Act5c </Td><Td> 40uL </Td></Tr> | ||
+ | <Tr><Td> 4 buffer(NEB) </Td><Td> 5uL </Td></Tr> | ||
+ | <Tr><td> XbaⅠ-HF(NEB) <td> 0.5uL </td></tr> | ||
+ | <Tr><Td> BamHⅠ(NEB )</Td><Td> 0.5uL </Td></Tr> | ||
+ | <Tr><Td> 100×BSA </Td><Td> 0.5uL </Td></Tr> | ||
+ | <Tr><Td> dH<sub>2</sub>O </Td><Td> 3.5uL </Td></Tr> | ||
+ | <Tr><Td> Total </Td><Td> 50uL </Td></Tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/2/27/0910akit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> G-1 and G-2 </Td><Td> G-3 and G-4 </Td></Tr> | ||
+ | <tr><td> DNA template </td><td> 1uL </td><Td> 4uL </Td></tr> | ||
+ | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td><Td> 5uL </Td></Tr> | ||
+ | <tr><td> 2mM dNTPs </td><td> 5uL </td><Td> 5uL </Td></tr><tr> | ||
+ | <td> 25mM MgSO4 </td><td> 1.6uL </td><Td> 1.6uL </Td></tr> | ||
+ | <tr><td> 10P 5’ primer </td><td> 1uL </td><Td> 1uL </Td></tr> | ||
+ | <tr><td> 10P 3’ primer </td><td> 1uL </td><Td> 1uL </Td></tr> | ||
+ | <tr><td> KOD plus </td><td> 1uL </td><Td> 1uL </Td></tr> | ||
+ | <tr><td> dH<sub>2</sub>O </td><td> 33.4uL </td><Td> 31.4uL </Td></tr> | ||
+ | <tr><td> Total </td><td> 50uL </td><Td> 50uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> | ||
+ | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> | ||
+ | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> | ||
+ | <tr><td> 57℃(G-1 and G-3) or 59℃(G-2 and G-4) </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> | ||
+ | <td> 68℃ </td><td> 2min10sec </td><Td> 30 cycle </Td></tr> | ||
+ | <tr><td> 68℃ </td><td> 2min10sec </td><Td></Td></tr> | ||
+ | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/e/eb/0910bkit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> pSB1C3(PCR) </Td><Td> 40uL </Td></Tr> | ||
+ | <tr><td> 4 buffer(NEB) </td><Td> 5uL </Td></tr> | ||
+ | <Tr><Td> EcoRⅠ-HF(NEB) </Td><Td> 0.5uL </Td></Tr> | ||
+ | <tr><td> SpeⅠ(NEB) </td><Td> 0.5uL </Td></tr><tr> | ||
+ | <td> 2100×BSA </td><Td> 0.5uL </Td></tr> | ||
+ | <tr><td> dH<sub>2</sub>O <Td> 3.5uL </Td></tr> | ||
+ | <tr><td> Total </td><Td> 50uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> 13uL </Td></Tr> | ||
+ | <tr><td> 3 buffer(NEB) </td><Td> 2uL </Td></tr> | ||
+ | <Tr><Td> BglⅡ(NEB) </Td><Td> 0.5uL </Td></Tr> | ||
+ | <tr><td> dH<sub>2</sub>O <Td> 2.5uL </Td></tr> | ||
+ | <tr><td> Total </td><Td> 20uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/8/86/0910ckit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> GAL4 </Td><Td> 20uL </Td></Tr> | ||
+ | <tr><td> 4 buffer(NEB) </td><Td> 4uL </Td></tr> | ||
+ | <Tr><Td> XbaⅠ(NEB) </Td><Td> 0.7uL </Td></Tr> | ||
+ | <Tr><Td> SpeⅠ(NEB) </Td><Td> 0.7uL </Td></Tr> | ||
+ | <Tr><Td> 100×BSA </Td><Td> 0.4uL </Td></Tr> | ||
+ | <tr><td> dH<sub>2</sub>O </td><Td> 14.6uL </Td></tr> | ||
+ | <tr><td> Total </td><Td> 40uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/3/39/0910dkit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> GAL4 </Td><Td> 40uL </Td></Tr> | ||
+ | <tr><td> 3 buffer(NEB) </td><Td> 5uL </Td></tr> | ||
+ | <Tr><Td> BglⅡ(NEB) </Td><Td> 0.5uL </Td></Tr> | ||
+ | <tr><td> dH<sub>2</sub>O </td><Td> 4.5uL </Td></tr> | ||
+ | <tr><td> Total </td><Td> 50uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> GAL4(Bgl Ⅱ cut) </Td><Td> 40uL </Td></Tr> | ||
+ | <tr><td> 4 buffer(NEB) </td><Td> 5uL </Td></tr> | ||
+ | <Tr><Td> SpeⅠ </Td><Td> 0.5uL </Td></Tr> | ||
+ | <Tr><Td> 100×BSA </Td><Td> 0.5uL </Td></Tr> | ||
+ | <tr><td> dH<sub>2</sub>O </td><Td> 4uL </Td></tr> | ||
+ | <tr><td> Total </td><Td> 50uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/6/6d/0910ekit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> 1uL </Td></Tr> | ||
+ | <tr><td></td><Td> 2uL </Td></tr> | ||
+ | <Tr><Td> EGFP or LacZ </Td><Td> 2uL </Td></Tr> | ||
+ | <tr><td> Ligation high </td><Td> 5uL </Td></tr> | ||
+ | <tr><td> Total </td><Td> 10uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> 1uL </Td></Tr> | ||
+ | <tr><td></td><Td> 2uL </Td></tr> | ||
+ | <Tr><Td></Td><Td> 2uL </Td></Tr> | ||
+ | <tr><td> Ligation high </td><Td> 5uL </Td></tr> | ||
+ | <tr><td> Total </td><Td> 10uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <h2>September 11th</h2> | ||
+ | <br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> pSB1C3-UAS-1, -2, -3, -4 </Td><Td> 5uL </Td></Tr> | ||
+ | <tr><td> 3buffer(NEB) </td><Td> 2uL </Td></tr> | ||
+ | <Tr><Td> BglⅡ </Td><Td> 0.5uL </Td></Tr> | ||
+ | <td> dH<sub>2</sub>O </td><Td> 12.5uL </Td></tr> | ||
+ | <td> Total </td><Td> 20uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/0/0c/0911akit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> GAL4 </Td><Td> 40uL </Td></Tr> | ||
+ | <tr><td> 4 buffer(NEB) </td><Td> 5uL </Td></tr> | ||
+ | <Tr><Td> XbaⅠ(NEB) </Td><Td> 0.5uL </Td></Tr> | ||
+ | <Tr><Td> SpeⅠ(NEB) </Td><Td> 0.5uL </Td></Tr><Tr> | ||
+ | <Td> CIP </Td><Td> 0.5uL </Td></Tr><Tr> | ||
+ | <Td> 100×BSA </Td><Td> 0.5uL </Td></Tr> | ||
+ | <td> dH<sub>2</sub>O </td><Td> 3uL </Td></tr> | ||
+ | <td> Total </td><Td> 40uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <img src="https://static.igem.org/mediawiki/2012/d/da/0911kit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> 1uL </Td></Tr> | ||
+ | <tr><td></td><Td> 2.5uL </Td></tr> | ||
+ | <Tr><Td></Td><Td> 2.5uL </Td></Tr> | ||
+ | <td> Ligation high </td><Td> 6uL </Td></tr> | ||
+ | <td> Total </td><Td> 12uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td></Td><Td> 2uL </Td></Tr> | ||
+ | <tr><td></td><Td> 2.5uL </Td></tr> | ||
+ | <Tr><Td></Td><Td> 1.5uL </Td></Tr> | ||
+ | <td> Ligation high </td><Td> 6uL </Td></tr> | ||
+ | <td> Total </td><Td> 12uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | |||
+ | <h2>September 12th</h2> | ||
+ | <br> | ||
+ | Composition | ||
+ | <br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> pSB1C3-UAS-1, -2 </Td><Td> 40uL </Td></Tr> | ||
+ | <tr><td> 3buffer(NEB) </td><Td> 5uL </Td></tr> | ||
+ | <Tr><Td> SpeⅠ </Td><Td> 1uL </Td></Tr> | ||
+ | <td> 100×BSA </td><Td> 0.5uL </Td></tr><tr> | ||
+ | <td> dH<sub>2</sub>O </td><Td> 3.5uL </Td></tr> | ||
+ | <td> Total </td><Td> 50uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | The digested DNA were applied to agarose gel electrophoresis. We isolated DNA from the gel by using QIA quick Gel Extraction Kit. | ||
+ | <br><br> | ||
+ | Photo of Agarose gel. | ||
+ | <br> | ||
+ | <img src="https://static.igem.org/mediawiki/2012/a/ab/0912kit.png" width="500" height="300"> | ||
+ | <br><br> | ||
+ | <strong>1-1-48 and 2-2-6 Ligation</strong> | ||
+ | <br> | ||
+ | DNA fragment carrying EGFP or LacZ was ligated into the BglII and SpeI digested pSB1C3-UAS DNA .for 1hour at 16℃. | ||
+ | <br><br> | ||
+ | Ligation reactions are listed below. | ||
+ | <br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> pSB1C3-UAS (double digested with BglII and SpeⅠ) </Td><Td> 1uL </Td></Tr> | ||
+ | <tr><td> EGFP fragments or LacZ fragments </td><Td> 2uL </Td></tr> | ||
+ | <Tr><Td> EGFP or LacZ </Td><Td> 2uL </Td></Tr><tr> | ||
+ | <td> Ligation high </td><Td> 2.5uL </Td></tr><tr> | ||
+ | <td> dH<sub>2</sub>O </td><Td> 2uL </Td></tr><tr> | ||
+ | <td> Total </td><Td> 7.5uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | We ligated DNA fragment carrying GAL4 and DNA fragments carrying HS promoter or Act5c promoter-enhancer to pSB1C3 DNA for 1hour at 16℃. | ||
+ | <br><br> | ||
+ | Composition | ||
+ | <br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td> pSB1C3 (PCR) (double digested with XbaⅠand SpeI) </Td><Td> 2uL </Td></Tr> | ||
+ | <tr><td> GAL4(double digested with BglⅡ and SpeⅠ) </td><Td> 2uL </Td></tr> | ||
+ | <Tr><Td> HS fragments or Act5c fragments (double digested with XbaⅠand BamHⅠ) </Td><Td> 2uL </Td></Tr><tr> | ||
+ | <td> Ligation high </td><Td> 6uL </Td></tr><tr> | ||
+ | <td> Total </td><Td> 12uL </Td></tr> | ||
+ | </Table> | ||
+ | <br><BR> | ||
+ | <strong>1-1-49 and 2-2-7 Transformation of E. coli by Ligation products</strong> | ||
+ | <br> | ||
+ | We did transformation of E. coli Xl-1 blue with each of ligation products. Finally, we spread them on the LB Chloramphenicol(+) plate and incubated for 16 hours at 37℃. | ||
+ | |||
+ | <br><br> | ||
+ | |||
+ | |||
+ | </div> | ||
+ | </div> | ||
+ | </table> | ||
+ | </td></tr> | ||
+ | |||
+ | </table> | ||
+ | </body> | ||
+ | |||
+ | |||
+ | |||
</html> | </html> |
Revision as of 10:12, 25 September 2012
September 6th
September 7th
September 8th
September 9th
September 10th
September 11th
September 12thComposition
The digested DNA were applied to agarose gel electrophoresis. We isolated DNA from the gel by using QIA quick Gel Extraction Kit. Photo of Agarose gel. 1-1-48 and 2-2-6 Ligation DNA fragment carrying EGFP or LacZ was ligated into the BglII and SpeI digested pSB1C3-UAS DNA .for 1hour at 16℃. Ligation reactions are listed below.
We ligated DNA fragment carrying GAL4 and DNA fragments carrying HS promoter or Act5c promoter-enhancer to pSB1C3 DNA for 1hour at 16℃. Composition
1-1-49 and 2-2-7 Transformation of E. coli by Ligation products We did transformation of E. coli Xl-1 blue with each of ligation products. Finally, we spread them on the LB Chloramphenicol(+) plate and incubated for 16 hours at 37℃. |