Team:UC Davis/notebook/initiator.html

From 2010.igem.org

(Difference between revisions)
Line 91: Line 91:
<li>It is most practical to cut RBS as a vector because RBS is less than 100 bp long as an insert, making it hard to see and extract on the gel.</li>
<li>It is most practical to cut RBS as a vector because RBS is less than 100 bp long as an insert, making it hard to see and extract on the gel.</li>
<li>The sticky ends from the XbaI and SpeI, when ligated together, will form an 8 bp scar.  Thus, XbaI and SpeI will no longer be able to cut here.</li>
<li>The sticky ends from the XbaI and SpeI, when ligated together, will form an 8 bp scar.  Thus, XbaI and SpeI will no longer be able to cut here.</li>
 +
</ul><br />
 +
 +
<a name="rbsc0051"><h1>'RBS & BBa_C0051' to Terminator</h1></a>
 +
<ul>
 +
<li><a href="https://2010.igem.org/Team:UC_Davis/protocols/culture.html" class="help">Cultured</a> cells that have parts: 'RBS & BBa_C0051', BBa_B0015</li>
 +
<li><a href="https://2010.igem.org/Team:UC_Davis/protocols/miniprep.html" class="help">Miniprepped</a> 'RBS & BBa_C0051' and BBa_B0015</li>
</ul>
</ul>
 +
 +
<table border="0" margin="1" width="650px" padding="5px">
 +
<tr>
 +
<td>
 +
    <table border="0" margin="1" width="320px" padding="5px">
 +
    <tr>
 +
      <td><img src="https://static.igem.org/mediawiki/2010/9/9a/Stopplasmid.jpg"></td>
 +
    </tr>
 +
    <tr>
 +
      <td><ul><li><a href="https://2010.igem.org/Team:UC_Davis/protocols/digestion.html" class="help">Digest</a> with EcoRI & XbaI</li></ul></td>
 +
    </tr>
 +
    <tr>
 +
      <td><img src="https://static.igem.org/mediawiki/2010/b/bb/RBSc0051.jpg"></td>
 +
    </tr>
 +
    <tr>
 +
      <td><ul>
 +
        <li><a href="https://2010.igem.org/Team:UC_Davis/protocols/gelelectrophoresis.html" class="help">Run on gel</a></li>
 +
        <li><a href="https://2010.igem.org/Team:UC_Davis/protocols/gelextraction.html" class="help">Gel extract</a></li></ul></td>
 +
    </tr>
 +
    <tr>
 +
      <td><img src="https://static.igem.org/mediawiki/2010/7/70/Xstopopenvector.jpg"></td>
 +
    </tr>
 +
    </table>
 +
</td>
 +
<td>
 +
  <table border="0" margin="1" width="320px" padding="5px">
 +
    <tr>
 +
      <td><img src="https://static.igem.org/mediawiki/2010/b/bb/RBSc0051.jpg"></td>
 +
    </tr>
 +
    <tr>
 +
      <td><ul><li><a href="https://2010.igem.org/Team:UC_Davis/protocols/digestion.html" class="help">Digest</a> with EcoRI & SpeI</li></ul> </td>
 +
    </tr>
 +
    <tr>
 +
      <td><img src="https://static.igem.org/mediawiki/2010/9/94/RBSc0051cut.jpg"></td>
 +
    </tr>
 +
    <tr>
 +
      <td><ul><li>Separate fragments through <a href="https://2010.igem.org/Team:UC_Davis/protocols/gelelectrophoresis.html" class="help"> gel electrophoresis</a></li><li><a href="https://2010.igem.org/Team:UC_Davis/protocols/gelextraction.html" class="help">Gel extract</a></li></ul></td>
 +
    </tr>
 +
    <tr>
 +
      <td><img src="https://static.igem.org/mediawiki/2010/d/de/RBSc0051insertx.jpg"></td>
 +
    </tr>
 +
  </table>
 +
</td>
 +
</tr>
 +
</table><p><br />
 +
 +
<ul>
 +
<li><a href="https://2010.igem.org/Team:UC_Davis/protocols/ligation.html" class="help">Ligate</a> the RBS and BBa_C0051 fragments together and <a href="https://2010.igem.org/Team:UC_Davis/protocols/transformation.html" class="help">transform</a> on carb plates.</li>
 +
<li>Depending on the outcome of visible colonies on the experimental and control plates, a <a href="https://2010.igem.org/Team:UC_Davis/protocols/pcrscreen.html" class="help">PCR screening</a> may be needed.
 +
</ul><br />
 +
 +
<img src="https://static.igem.org/mediawiki/2010/e/eb/RBSc0051stop.jpg"><br />
                            
                            
         </td>
         </td>

Revision as of 10:12, 19 September 2010

The Initiator

Starting Up

  • Hydrated parts: BBa_B0034, BBa_C0051, BBa_B0015, BBa_R0082.
  • Transformed these parts into DH5α competent cells.


RBS to BBa_C0051


  • Ligate the RBS and BBa_C0051 fragments together and transform on carb plates.
  • Depending on the outcome of visible colonies on the experimental and control plates, a PCR screening may be needed.


Assembly notes
  • It is most practical to cut RBS as a vector because RBS is less than 100 bp long as an insert, making it hard to see and extract on the gel.
  • The sticky ends from the XbaI and SpeI, when ligated together, will form an 8 bp scar. Thus, XbaI and SpeI will no longer be able to cut here.

'RBS & BBa_C0051' to Terminator
  • Cultured cells that have parts: 'RBS & BBa_C0051', BBa_B0015
  • Miniprepped 'RBS & BBa_C0051' and BBa_B0015


  • Ligate the RBS and BBa_C0051 fragments together and transform on carb plates.
  • Depending on the outcome of visible colonies on the experimental and control plates, a PCR screening may be needed.


We would like to take a moment to thank all of our sponsors for their very generous donations, as we could not have done this without your help!

We would also like to thank and acknowledge:
Our Advisors
Marc Facciotti
Ilias Tagkopoulos
Technical Guidance
David Larsen
Andrew Yao
Visiting iGEMer
Jia Li of Zhejiang University (TEAM ZJU-China)
cI Promoter Screen
Drew Endy - Stanford
Thomas Schneider - NIH
Want to sponsor us? Send an email to mtfacciotti@ucdavis.edu to discuss various ways you can help! :)

Retrieved from "http://2010.igem.org/Team:UC_Davis/notebook/initiator.html"