Team:Washington/Notebook/colony PCR
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Latest revision as of 05:12, 15 October 2009
Colony PCR
- Prepare one sterile 0.6mL tube with the following reaction mixture for each colony you intend to pick.
- 5uL Qiagen Master Mix
- 1uL 40uM VF2
- 1uL 40uM VR
- Prepare one sterile 0.6mL tube with 20uL sterile diH2O for each colony you intend to pick.
- Pick colonies
- Pick a single colony using a micropipettor with sterile tip. The pippettor should be set to 3uL
- Aspirate colony into 20uL diH2O vigorously to transfer cells to diH2O
- Transfer 3uL of diH2O containing cells to reaction mixture set up in step 1
- Run reaction
- 94C - 3min
- 94C - 30s
- 55C - 30s
- 72C - 1min / kb gene
- repeat 2 - 4 29x
- 72C - 10min
- 4C - forever