Team:KIT-Kyoto/kazukokekokko
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{{KIT-Kyoto}} | {{KIT-Kyoto}} | ||
+ | {{KIT-Kyoto.Header}} | ||
<html> | <html> | ||
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+ | <meta charset="UTF-8"> | ||
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<style type="text/css"> | <style type="text/css"> | ||
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+ | <table border="0" width="965px" align="center"><tr><td width="165px" valign="top" | ||
- | + | align="left"><div id="HIDARI"> | |
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<br> | <br> | ||
- | < | + | <div> |
- | < | + | <ul class="navi"> |
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- | + | <ul class="menu"> | |
- | + | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-week1"><img src="https://static.igem.org/mediawiki/2012/1/1b/Side_week1kit.jpg" width="150" height="30"></a></li> | |
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- | + | <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> | |
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- | + | <div class="category"><img src="https://static.igem.org/mediawiki/2012/6/61/Side_meetingkit.jpg" width="150" height="30"></div> | |
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- | + | <li><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-Meeting-may"><img src="https://static.igem.org/mediawiki/2012/5/5d/Side_maykit.jpg" width="150" height="30"></a></li> | |
- | + | <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> | |
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- | + | <li> | |
- | + | <div class="category"><a href="https://2012.igem.org/Team:KIT-Kyoto/Notebook-Design"><img src="https://static.igem.org/mediawiki/2012/8/8f/Side_designnotekit.jpg" width="150" height="30"></a></div> | |
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</div> | </div> | ||
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<br> | <br> | ||
- | + | </td> | |
+ | <td width="800px" height="510px" valign="top"> | ||
+ | <div id="MIGI"> | ||
+ | <font color="#f5b1aa"><u>TNFAIP3</u></font> | ||
<br> | <br> | ||
- | + | <font color="#f5b1aa"><u>PARTS</u></font> | |
<br> | <br> | ||
- | < | + | <h2>BP reaction by Invitrogen gateway system</h2> |
<br> | <br> | ||
+ | 1. PCR using primers containing the attB sequence. | ||
+ | <br> | ||
+ | 2. Purify PCR product. | ||
+ | <br> | ||
+ | 3. Add the following components to a 1.5 mL microcentrifuge tube at room temperature and mix: | ||
+ | <br><br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td>attB PCR product</Td><Td>75 ng/reaction (1-7 μL)</Td></Tr> | ||
+ | <Tr><Td>pDONR vector</Td><Td>150ng/reaction (1 μL)</Td></Tr> | ||
+ | <Tr><Td>TE Buffer</Td><Td>to 8 μL</Td></Tr> | ||
+ | </Table> | ||
+ | <br> | ||
+ | 4. Vortex BP ClonaseII enzyme mix briefly. Add 1 - 2 μL to the components above and mix well by vortexing and spin them down. | ||
+ | <br> | ||
+ | 5. Incubate reaction at 25˚C for more than 1 hour. | ||
+ | <br> | ||
+ | 6. Add 1 μL of 2 μg/μL Proteinase K solution and vortex briefly. | ||
+ | <br> | ||
+ | 7. Incubate at 37˚C for 10 minutes. | ||
+ | <br> | ||
+ | <br> | ||
+ | <h2>LR reaction by Invitrogen gateway system</h2> | ||
+ | <br> | ||
+ | 1. Add the following components to a 1.5 mL microcentrifuge tube at room temperature and mix: | ||
+ | <br><br> | ||
+ | <Table Border Cellspacing="0"> | ||
+ | <Tr><Td>Entry clones</Td><Td>50-150 ng/reaction (1-7 μL)</Td></Tr> | ||
+ | <tr><td>Destination vector</td><td>150ng/reaction (1 μL)</td></tr> | ||
+ | <Tr><Td>TE Buffer</Td><Td>to 8 − 9 μL</Td></Tr> | ||
+ | </Table> | ||
+ | <br> | ||
+ | 2. Vortex LR ClonaseII enzyme mix briefly. Add 1 – 2 μL, to the components above and mix well by vortexing and spin down. | ||
+ | <br> | ||
+ | 3. Incubate reaction at 25˚C for 16 hours. | ||
+ | <br> | ||
+ | 4. Add 1 μL of 2 μg/μL Proteinase K solution and vortex briefly. | ||
+ | <br> | ||
+ | <br> | ||
+ | 5. Incubate at 37˚C for 10 minutes. | ||
+ | <br> | ||
+ | <br> | ||
+ | </div> | ||
+ | </td> | ||
+ | </tr> | ||
- | + | <tr> | |
+ | <td ColSpan="2" width="965"> | ||
+ | <div id="NAKAMI"> | ||
+ | <br> | ||
+ | <div align="center"> | ||
+ | <strong>Our Sponsors</strong> | ||
+ | <br><br> | ||
+ | <a href="http://www.kit.ac.jp/"><img src="https://static.igem.org/mediawiki/2012/e/ea/KITkyoto.png" width="100" height="100"></a> | ||
+ | <br> | ||
+ | <br> | ||
</div> | </div> | ||
</div> | </div> | ||
- | + | </td> | |
- | </div> | + | </tr> |
- | + | </div> | |
+ | </table> | ||
</body> | </body> | ||
</html> | </html> |
Latest revision as of 12:49, 25 September 2012
TNFAIP3
PARTS BP reaction by Invitrogen gateway system1. PCR using primers containing the attB sequence. 2. Purify PCR product. 3. Add the following components to a 1.5 mL microcentrifuge tube at room temperature and mix:
4. Vortex BP ClonaseII enzyme mix briefly. Add 1 - 2 μL to the components above and mix well by vortexing and spin them down. 5. Incubate reaction at 25˚C for more than 1 hour. 6. Add 1 μL of 2 μg/μL Proteinase K solution and vortex briefly. 7. Incubate at 37˚C for 10 minutes. LR reaction by Invitrogen gateway system1. Add the following components to a 1.5 mL microcentrifuge tube at room temperature and mix:
2. Vortex LR ClonaseII enzyme mix briefly. Add 1 – 2 μL, to the components above and mix well by vortexing and spin down. 3. Incubate reaction at 25˚C for 16 hours. 4. Add 1 μL of 2 μg/μL Proteinase K solution and vortex briefly. 5. Incubate at 37˚C for 10 minutes. |
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