Team:KIT-Kyoto/kazukokekokko

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<table border="0" width="965px" align="center"><tr><td width="165px" valign="top"
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<font size="5">pre page</font>
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align="left"><div id="HIDARI">
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<img src="" align="left" width="150" height="150">NAME
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<img src="" align="right" width="150" height="150">NAME
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            <div class="category"><img src="https://static.igem.org/mediawiki/2012/1/1f/Side_labnotekit.jpg" width="150" height="30"></div>
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                <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-week2"><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-week3"><img src="https://static.igem.org/mediawiki/2012/e/e0/Side_week3kit.jpg" width="150" height="30"></a></li>
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<br>
<br>
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pfofile ex.<A href="https://2012.igem.org/Team:KIT-Kyoto/Team-Kazuko" >いちいち押すやつ</A>
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</td>
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<td width="800px" height="510px" valign="top">
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<div id="MIGI">
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<font color="#f5b1aa"><u>TNFAIP3</u></font>
<br>
<br>
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MENU 参考 <A href="https://2010.igem.org/wiki/index.php?title=Template:KIT-Kyoto/menu10&action=edit">go</A>
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<font color="#f5b1aa"><u>PARTS</u></font>
<br>
<br>
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見出し <a href="http://midashi-maker.v-colors.com/">go</a>
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<h2>BP reaction by Invitrogen gateway system</h2>
<br>
<br>
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<a href="http://www.colordic.org/w/"> 日本の伝統色</a>
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1. PCR using primers containing the attB sequence.
<br>
<br>
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候補<a href="https://2012.igem.org/Team:KIT-Kyoto/01wiki"> go </a>
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2. Purify PCR product.
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<br>
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3. Add the following components to a 1.5 mL microcentrifuge tube at room temperature and mix:
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<br><br>
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<Table Border Cellspacing="0">
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<Tr><Td>attB PCR product</Td><Td>75 ng/reaction (1-7 μL)</Td></Tr>
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<Tr><Td>pDONR vector</Td><Td>150ng/reaction (1 μL)</Td></Tr>
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<Tr><Td>TE Buffer</Td><Td>to 8 μL</Td></Tr>
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</Table>
 +
<br>
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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>
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5. Incubate reaction at 25˚C for more than 1 hour.
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<br>
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6. Add 1 μL of 2 μg/μL Proteinase K solution and vortex briefly.
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<br>
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7. Incubate at 37˚C for 10 minutes.
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<br>
 +
<br>
 +
<h2>LR reaction by Invitrogen gateway system</h2>
 +
<br>
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1. Add the following components to a 1.5 mL microcentrifuge tube at room temperature and mix:
 +
<br><br>
 +
<Table Border Cellspacing="0">
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<Tr><Td>Entry clones</Td><Td>50-150 ng/reaction (1-7 μL)</Td></Tr>
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<tr><td>Destination vector</td><td>150ng/reaction (1 μL)</td></tr>
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<Tr><Td>TE Buffer</Td><Td>to 8 − 9 μL</Td></Tr>
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</Table>
 +
<br>
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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>
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</div>
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</td>
 +
</tr>
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<tr>
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<td ColSpan="2" width="965">
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<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>
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<br>
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<br>
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</div>
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</td>
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Latest revision as of 12:49, 25 September 2012
TNFAIP3
PARTS

BP reaction by Invitrogen gateway system


1. 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:

attB PCR product75 ng/reaction (1-7 μL)
pDONR vector150ng/reaction (1 μL)
TE Bufferto 8 μL

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 system


1. Add the following components to a 1.5 mL microcentrifuge tube at room temperature and mix:

Entry clones50-150 ng/reaction (1-7 μL)
Destination vector150ng/reaction (1 μL)
TE Bufferto 8 − 9 μL

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.


Our Sponsors



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