IGEM:Groningen/Notebook/iGEM 2011/2011/07/15: Difference between revisions

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==Entry title==
==15-07-11==
* Insert content here...
<br>
<br> Plates of yesterday's transformation looks better than transformations before. Self ligation plates contain 5 to 10 times less <br> colonies than the rest of the plates.
<br> Colony PCR was done as followed:
<br> Mastermix scheme:
<br> Taq 10× buffer with NH4SO4 without MgCl2: 58.00μl
<br> dNTPs 10mM each: 11.6μl
<br> MgCl2: 34.8μl
<br> Taq polymerase 5u/μl: 2.90μl
<br> Biobrick vector forward primer 10μM: 11.6μl
<br> Biobrick vector reverse primer 10μM: 11.6μl
<br> MilliQ water: 449.50μl
<br>
<br> PCR conditions:
<br> Preheated lid: 111°C
<br> Denaturation: 94°C for 10 min.
<br> Cycle (33×)
<br>  Denaturation: 94°C for 30s.
<br>  Annealing: 60°C for 30s.
<br>  Extension: 72°C for 2 min.
<br> Final extension: 72°C for 10 min.
<br> Store at 4°C infinite
<br>
<br> PCR products were analysed on a 1% agarose gel with TBE.
<br>




Revision as of 01:31, 15 July 2011

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15-07-11



Plates of yesterday's transformation looks better than transformations before. Self ligation plates contain 5 to 10 times less
colonies than the rest of the plates.
Colony PCR was done as followed:
Mastermix scheme:
Taq 10× buffer with NH4SO4 without MgCl2: 58.00μl
dNTPs 10mM each: 11.6μl
MgCl2: 34.8μl
Taq polymerase 5u/μl: 2.90μl
Biobrick vector forward primer 10μM: 11.6μl
Biobrick vector reverse primer 10μM: 11.6μl
MilliQ water: 449.50μl

PCR conditions:
Preheated lid: 111°C
Denaturation: 94°C for 10 min.
Cycle (33×)
Denaturation: 94°C for 30s.
Annealing: 60°C for 30s.
Extension: 72°C for 2 min.
Final extension: 72°C for 10 min.
Store at 4°C infinite

PCR products were analysed on a 1% agarose gel with TBE.


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