Team:Groningen/Template/MODULE/Notebook/characterisation/week6

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Testing the constructs with purified P2, Plas, J23101 and J23115, Inserting the constructs (with promotor collection) in PIL vector
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Testing the constructs RBS-NisA-double terminator (BBa_K1365556) and RBS-Superfolded GFP-double terminator (BBa_K1365555) with some different promoter: P2, PLas, and two constitutive promoters (J23101 and J23115), and inserting these constructs in pIL253A vector
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<div class="item">Overnight ligation , transformation and colony PCR</div>
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<div class="item">The ligations of the different constructs were performed overnight, after which the ligated plasmids were transformed to <i>L. lactis</i> NZ9000, and finally a colony PCR was performed with several colonies of each construct</div>
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<div class="item">P2 and Plas did not show any bands on the gel so proceeded with J23101 and J23115</div>
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<div class="item">The construct with the P2 and PLas did not show any bands on the gel, therefore further steps were only done with the constructs containing the two constitutive promoters</div>
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<div class="item">The constructs with the promotor collection was subjected to ligation, but this time PIL vector was used and transformed in competent Lactococcus lactis</div>
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<div class="item">The same constructs with the promoters from the promoter collection of Uppsala (see September 19 till October 5) were subjected to ligation again, but this time the vector pIL235A was used instead of pSB1K3</div>
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<div class="item">The transformed cells were plated on M17 plates with eyrthromycin</div>
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<div class="item">These constructs were transformed to <i>L. lactis</i> NZ9000, and plated on M17 agar with erythromycin</div>
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Revision as of 17:35, 17 October 2014

6 - 12 October
 
Testing the constructs RBS-NisA-double terminator (BBa_K1365556) and RBS-Superfolded GFP-double terminator (BBa_K1365555) with some different promoter: P2, PLas, and two constitutive promoters (J23101 and J23115), and inserting these constructs in pIL253A vector
 
The ligations of the different constructs were performed overnight, after which the ligated plasmids were transformed to L. lactis NZ9000, and finally a colony PCR was performed with several colonies of each construct
The construct with the P2 and PLas did not show any bands on the gel, therefore further steps were only done with the constructs containing the two constitutive promoters
The same constructs with the promoters from the promoter collection of Uppsala (see September 19 till October 5) were subjected to ligation again, but this time the vector pIL235A was used instead of pSB1K3
These constructs were transformed to L. lactis NZ9000, and plated on M17 agar with erythromycin
 
 
 
 
 

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