Team:Paris Saclay/Notebook/September/4
From 2014.igem.org
Contents |
Thursday 4st September
Lab Work
By Mélanie
Lemon Scent
Verfication of the pPS3/4/5 plasmid
Due to the strange results obtained last day, I do a PCR of the insert with the differents plasmid : I use the same PCR condition than September 2 but with the right primer and plasmid
We can see that we don't have any insert in our plasmid
PCR verification
We find some other PCR of BBa K762100 and GS and BBa_K517003 so we check it by electrophoresis
And I purify the PCR
Well 1 = Ladder
Well 2 = GS
Well 3 = PS
Well 4 = CAD
We can see that we don't have lost a lot of our PCR product
cloning
Cloning of PCR purification in a TA plasmid (Topo plasmid)
First I had to add AAA to the PCR
component | volume |
---|---|
buffer | 1μl |
dATP | 1μl |
PCR | 7.5μl |
Taq | 0.5μl |
72° 15'
and
component | volume |
---|---|
H2O | 1μl |
buffer | 1μl |
vector | 1μl |
cloning | 1μl |
30' at room temperature
Transformation
Transformation with competent bacteria
add DNA (Top vector)
30' on ice 45' 42°c 30' at 37°
spread on petri dish