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

 
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<h4>Transforming E. Coli DH5a to amplify pKD3 for pRED/ET engineering</h4>
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<h4>Transforming E. Coli DH5a to Amplify pKD3 for pRED/ET Engineering</h4>
 
<em>2018/05/28</em>
 
<em>2018/05/28</em>
 
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       <td>Protocol:</td>
 
       <td>Protocol:</td>
 
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<a href="https://static.igem.org/mediawiki/2018/c/ca/T--Munich--WL1_Electrocompetent_transformation.pdf" target="_blank">Electrocompetent transformation</a>
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<a href="https://static.igem.org/mediawiki/2018/c/ca/T--Munich--WL1_Electrocompetent_transformation.pdf" target="_blank">Electro-transformation</a>
 
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       <td>Notes:</td>
 
       <td>Notes:</td>
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       <td>we also inoculated LB media blindly.</td>
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       <td>No notes.</td>
 
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       <td>Results:</td>
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       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
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       <td>no colonies, no growth; <br>
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       <td>No colonies, no growth; <br>
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We decided to use a resistance cassette from pSB1C3 without FRT sites
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We decided to use a resistance cassette from pSB1C3 without FRT sites.
 
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<h4>Amplifying a selection cassette from pSB1C3</h4>
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<h4>Amplifying a Selection Cassette from pSB1C3</h4>
 
<em>2018/05/29</em>
 
<em>2018/05/29</em>
 
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       <td>Notes:</td>
 
       <td>Notes:</td>
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       <td>Primer with 50bp homology regions ????, TA: 67°C, Elongation: 45s</td>
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       <td>PCR with two primer pairs were tested to amplify a chloramphenicol resistance cassette for the knockout. Both pairs have a 50 bp overhang homology region to the knockout target in E.Coli genomes. One pair has longer annealing sequence (CAT_RecBCD-KO_5HA_long_rv  and CAT_RecBCD-KO_5HA_long_fw) and the other pair shorter. (CAT_RecBCD-KO_5HA_short_rv  and CAT_RecBCD-KO_5HA_short_fw).</td>
 
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       <td>Results:</td>
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       <td><a href="https://2018.igem.org/Team:Munich/Results" target="_blank">Results:</a></td>
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       <td>PIC. we thought the band might be what we wanted but because of how few DNA the extraction yielded, these samples were not used in further experiments.
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       <td> We thought the band might be what we wanted but because of how few DNA the extraction yielded, these samples were not used in further experiments.
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<figure class="figure">
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  <img src="https://static.igem.org/mediawiki/2018/b/bd/T--munich--20180605_CAT-KO.jpg" alt=" ">
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    <figcaption class="figure-caption"></figcaption>
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Latest revision as of 00:13, 18 October 2018

Transforming E. Coli DH5a to Amplify pKD3 for pRED/ET Engineering

2018/05/28
Participants: Dominic Schwarz
Protocol: Electro-transformation
Notes: No notes.
Results: No colonies, no growth;
We decided to use a resistance cassette from pSB1C3 without FRT sites.

Amplifying a Selection Cassette from pSB1C3

2018/05/29
Participants: Enikö Baligács
Protocol: PCR, Agarose gel, Gel extraction
Notes: PCR with two primer pairs were tested to amplify a chloramphenicol resistance cassette for the knockout. Both pairs have a 50 bp overhang homology region to the knockout target in E.Coli genomes. One pair has longer annealing sequence (CAT_RecBCD-KO_5HA_long_rv and CAT_RecBCD-KO_5HA_long_fw) and the other pair shorter. (CAT_RecBCD-KO_5HA_short_rv and CAT_RecBCD-KO_5HA_short_fw).
Results: We thought the band might be what we wanted but because of how few DNA the extraction yielded, these samples were not used in further experiments.