Team:Evry/Notebook/Transposons/13.10.14

From 2014.igem.org

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== BBa_1413044 retrieval of the transposon plasmid==
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<h4> BBa_1413044 retrieval of the transposon plasmid </h4>
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strategy:</p>
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<div align="center">
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<br><img src="https://static.igem.org/mediawiki/2014/3/33/Evry_Strat1.jpg" width="75%"/></div>
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<p>
Digestion of BBa_1413044 by BglII :
Digestion of BBa_1413044 by BglII :
<br>50 ul final volume, 5 ul Buffer NeB 3.1, 5 ul (20 ng/ul) BBa_1413044 in pSB1C3, 1 ul BglII, 39 ul H2O.
<br>50 ul final volume, 5 ul Buffer NeB 3.1, 5 ul (20 ng/ul) BBa_1413044 in pSB1C3, 1 ul BglII, 39 ul H2O.
<br>
<br>
Gel electrophoresis:
Gel electrophoresis:
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GEEEEEEEEEEEEEEEEEEEEEELLLLLLLLLL
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<img src="https://static.igem.org/mediawiki/2014/3/31/Evry_Bba_transpo.JPG" width="25%"/></div>
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<img src="" width="50%"/>
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<br>
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Purification sur gel:
Purification sur gel:
<br>
<br>
Kit thermoscientific "Gel extraction". Elution final in 30 ul.
Kit thermoscientific "Gel extraction". Elution final in 30 ul.
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<br></p>
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== Obtaining the Kan gene on pNK2 ==
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<h4>Obtaining the Kan gene on pNK2</h4>
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<br>
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<br><p>
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PCR: enzyme: Q5; template: 150 ng pNK2; oligo: F 82/ R 83; Tm tested: 55/60/65; elongation time: 1m30s
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PCR: enzyme: Q5; template: 150 ng pNK2 first three well from new stock, next ones the old stock; oligo: F 82/ R 83; Tm tested: 55/60/65; elongation time: 1m30s
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<img src="" width="50%"/>
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<img src="https://static.igem.org/mediawiki/2014/9/90/Evry_Kan_extract.JPG" width="40%"/></div>
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<br>
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<br><p>
Purification sur gel:
Purification sur gel:
<br>
<br>
Kit thermoscientific "Gel extraction". Elution final in 30 ul.
Kit thermoscientific "Gel extraction". Elution final in 30 ul.
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Latest revision as of 22:41, 17 October 2014

Picture

BBa_1413044 retrieval of the transposon plasmid


strategy:



Digestion of BBa_1413044 by BglII :
50 ul final volume, 5 ul Buffer NeB 3.1, 5 ul (20 ng/ul) BBa_1413044 in pSB1C3, 1 ul BglII, 39 ul H2O.
Gel electrophoresis:


Purification sur gel:
Kit thermoscientific "Gel extraction". Elution final in 30 ul.

Obtaining the Kan gene on pNK2


PCR: enzyme: Q5; template: 150 ng pNK2 first three well from new stock, next ones the old stock; oligo: F 82/ R 83; Tm tested: 55/60/65; elongation time: 1m30s


Purification sur gel:
Kit thermoscientific "Gel extraction". Elution final in 30 ul.

Oct 13