Difference between revisions of "Team:Munich/engineering2.html"

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<html>
 
<div class="event-info">
 
<div class="event-info">
  
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     </tr>
 
     </tr>
 
</table>
 
</table>
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<h4>Creating a selection cassette from pSB1C3</h4>
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<em>2018/08/30</em>
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<table class="table table-borderless">
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    <tr>
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      <td>Participants:</td>
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      <td>Dominic</td>
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    </tr>
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    <tr>
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      <td>Protocol:</td>
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      <td>restrictiondigest, PCR-purification</td>
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    </tr>
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    <tr>
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      <td>Notes:</td>
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      <td>DpnI
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</td>
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    </tr>
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<tr>
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      <td>Results:</td>
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      <td>CAP_recBCD 18ng/ul
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CAP_msbB 12ng/ul
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</td>
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    </tr>
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</table>
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<h4>pRED/ET genome engineering of delta msb-B and delta recBCD strains</h4>
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<em>2018/09/01</em>
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<table class="table table-borderless">
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    <tr>
 +
 +
      <td>Participants:</td>
 +
      <td>Dominic</td>
 +
    </tr>
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    <tr>
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      <td>Protocol:</td>
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      <td>pRED/ET engineering protocol</td>
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    </tr>
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    <tr>
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      <td>Notes:</td>
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      <td>the template for the resistance cassette for deletions was taken from a plasmid containing mRFP
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</td>
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    </tr>
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<tr>
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      <td>Results:</td>
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      <td>colonies on both plates
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 +
</td>
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    </tr>
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</table>
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<h4>Verifying deletion strains of E. Coli MG1655</h4>
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<em>2018/09/05</em>
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<table class="table table-borderless">
 +
    <tr>
 +
 +
      <td>Participants:</td>
 +
      <td>Dominic</td>
 +
    </tr>
 +
    <tr>
 +
      <td>Protocol:</td>
 +
      <td>Colony PCR, Agarose gel</td>
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    </tr>
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    <tr>
 +
      <td>Notes:</td>
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      <td>Primers: VF2, geno_msb-B_rv, geno_recBCD_rv;
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Ta: 48°C
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t= 1kb/min <br>
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RecBCD: 1,2,3,4,5 <br>
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msbB: 35,36,37,38,39
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expected: RecBCD 480bp
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Expected: msb-B 520bp
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</td>
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    </tr>
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<tr>
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      <td>Results:</td>
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      <td>all 5 picked colonies were positive for the insertion of the selection cassette PIC
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</td>
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    </tr>
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</table>
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<figure class="figure">
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  <img src="https://static.igem.org/mediawiki/2018/b/b3/T--Munich--Labbook_20180905_geno_analgel_recBCD12345%28443kb%29_msbB678910%28574kb%29.jpeg" class="figure-img img-fluid rounded" alt="awesome gel.">
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    <figcaption class="figure-caption">sick caption</figcaption>
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    </figure>
  
 
</div>
 
</div>
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</html>

Revision as of 15:17, 29 September 2018

Transforming E. Coli Rosetta and MG1655 for pRED/ET engineering

2018/08/28
Participants: Dominic
Protocol: epo trafo
Notes: Genetic engineering is planned in E. Coli Rosetta, MG1655 is E.Coli WT as backup.
Results: colonies only on E. Coli MG1655 plate; No colonies on E. Coli Rosetta

pRED/ET genome engineering of delta msb-B and delta recBCD strains

2018/08/29
Participants: Dominic
Protocol: pRED/ET engineering protocol
Notes: the template for the resistance cassette for deletions was taken from a plasmid containing mRFP
Results: red colonies. Elena (Advisor) and Dominic decided to repeat the experiment but to do a dpnI digest before to get rid of the template DNA.

Transforming E. Coli DH5a to find a reason for the contamination

2018/08/30
Participants: Dominic
Protocol: epo trafo
Notes: CAP_recBCD, CAP_msbb, psb1c3_mrfp in Dh5a
Results: red colonies on all plates -> mRFP contamination

Creating a selection cassette from pSB1C3

2018/08/30
Participants: Dominic
Protocol: restrictiondigest, PCR-purification
Notes: DpnI
Results: CAP_recBCD 18ng/ul CAP_msbB 12ng/ul

pRED/ET genome engineering of delta msb-B and delta recBCD strains

2018/09/01
Participants: Dominic
Protocol: pRED/ET engineering protocol
Notes: the template for the resistance cassette for deletions was taken from a plasmid containing mRFP
Results: colonies on both plates

Verifying deletion strains of E. Coli MG1655

2018/09/05
Participants: Dominic
Protocol: Colony PCR, Agarose gel
Notes: Primers: VF2, geno_msb-B_rv, geno_recBCD_rv; Ta: 48°C t= 1kb/min
RecBCD: 1,2,3,4,5
msbB: 35,36,37,38,39 expected: RecBCD 480bp Expected: msb-B 520bp
Results: all 5 picked colonies were positive for the insertion of the selection cassette PIC
awesome gel.
sick caption