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- Remove bacterial plates from refrigerator and put
pre-determined number of plates with the appropriate antibiotic into a 37oC
incubator.
- Thaw competent cells on ice. It is essential that cells
be completely thawed before pipetting. When cells are completely thawed, you
should see clearly two phases: the upper clear phase and the bottom cloudy
phase. Pipet slowly and don’t pipet cells up and down.
- Transfer 100 or 50 ml
of cells to a prechilled microcentrifuge tube or Falcon 352059 tube, add the DNA of
your choice (pure DNA or ligation mix) to the cells and incubate on ice for 5
minutes.
- Plate out desired amount on a prewarmed (37°C)
plate with the appropriate antibiotic and incubate in a 37°C
incubator. The plates should have been in the incubator for at least 30 min.
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