Team:Yale/Notebook/Week8

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<h1>Week 8: July 3-10, 2011</h1>
<h1>Week 8: July 3-10, 2011</h1>
<ul>
<ul>
-
<li>Run colony PCR on ZeAFP = successful; send in to Keck for sequencing verification</li>
+
<li>Designed and ordered primers with BioBrick prefix and suffix overhangs to PCR amplify following constructs <ul>
 +
<li>RiAFP-His</li>
 +
<li>eGFP-TEV-RiAFP-His</li>
 +
<li>eGFP-TmAFP-His</li>
 +
<li>eGFP-TEV-TmAFP-His</li>
 +
<li>TmAFP-His</li>
 +
</ul>
 +
<li>Set up PCR amplification with Phusion polymerase of aforementioned constructs, verified successful amplification: (show figure)</li>
 +
<li>Digest PCR fragments using XbaI and PstI, digest BioBrick BBa_I712074 (contains T7 polymerase) with SpeI and PstI</li>
 +
<li>Ligate using T4 ligase, transform cells and plate on Amp plates (backbone of plasmid is AmpR and KanR)</li>
 +
<li>Verification of successful ligation using colony PCR</li>
</ul>
</ul>
</div>
</div>
<div style="clear:both"></div>
<div style="clear:both"></div>
</div>
</div>

Latest revision as of 01:17, 28 September 2011

iGEM Yale

Week 8: July 3-10, 2011

  • Designed and ordered primers with BioBrick prefix and suffix overhangs to PCR amplify following constructs
    • RiAFP-His
    • eGFP-TEV-RiAFP-His
    • eGFP-TmAFP-His
    • eGFP-TEV-TmAFP-His
    • TmAFP-His
  • Set up PCR amplification with Phusion polymerase of aforementioned constructs, verified successful amplification: (show figure)
  • Digest PCR fragments using XbaI and PstI, digest BioBrick BBa_I712074 (contains T7 polymerase) with SpeI and PstI
  • Ligate using T4 ligase, transform cells and plate on Amp plates (backbone of plasmid is AmpR and KanR)
  • Verification of successful ligation using colony PCR

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