Team:Cambridge/Protocols/PCRcolony

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Colony PCR:

Risk Assessment


Used to amplify DNA directly from cell culture, the reaction is the same as for normal PCR but with modified reaction composition and cycle settings, here shown for a 50 µl reaction using Taq polymerase.

ReagentVolume (µl)Final Concentration
Water35.7
10 mM dNTPs1200 µM
10 x NH4 buffer51x
Forward Primer2.50.5 µM
Reverse Primer2.50.5 µM
Template Cells1.3 (from liquid culture or picked colony)
Taq polymerase 5u/µl10.1 u/ µl

Please refer to the standard PCR protocol for the remainder of this protocol.


Notes on colony PCR

  • This technique should only really be used for crude PCR assays, such as diagnostic PCR. If high quality DNA is desired, Miniprep followed by standard PCR should be used.
  • If taking a colony from a plate, resuspend the cells in ~50μl of water. Take 1 μl of this resuspension and use it as your template. Failing to do so will result in you having too much genomic DNA in your reaction, which will interfere with gel electrophoresis - you can tell if this has happened if a bright band appears where your wells are on the gel.
  • Resuspended cells can be stored in the fridge for a couple of days for eventual plating, if colony PCR proves successful.




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