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From 2011.igem.org

Contents 1 Wednesday 1.1 Lab Work 1.1.1 Plant CYPs 1.1.2 Human CYPs

Wednesday

Colonies on A1 - Finally. And our supervisors say that this is the easiest to work with. Hmm.

We still havent heard anything from our nice and helpful benefactors who supplied us with the human CYPs about the sequence and the composition of the plasmid. But we can't wayt any longer so we choose to believe that the plasmid contain ammphicilin. And then well get it sequenced - for this we will use the standard sequence of the middle of the CYPs which has most likely not been altered to much for expression in E.coli.

Lab Work

Plant CYPs

BioBrick A1

• We took the A1's out and put it in and overnight culture

BioBrick A2

• We purified A2 (second edition) with Mini prep from 2 different colonies.
• We analyzed A2 cut with Pst1 (which cut at the restrictionsite we want to remove)on an agarose gel. Sent them both to be sequences.

BioBrick B1

• Double digestion of B1 using the iGEM approach.
• Ligation of the BioBrick (CYP79 B1) and the plasmid backbone (pSB1C3).
• Transformation of the constructed plasmid into competent cells.

Human CYPs

• Transformed XL1-Blue with human cyp 3A4, 2C9, 1A1, 1A2 and hNRP.

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