Koeris/Notebook/2007-1-8

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Chemically competent bacterial cells

All cultures are grown at 37 deg C and 300 rpm shaking. The OD600 of the culture should be between 0.4 - 0.6. The culture is then centrifuged down at 3000rpm for 15min and the pellet resuspended in 1/10 volume of TSS. All steps are to be performed on ice. The tubes for the aliquots are also to be pre-chilled.

Reagents needed

The following reagents are needed for the protocol.

TSS (Transformation and storage solution)

For 50ml use the following ratios:

  • LB: 30ml
  • PEG: 5.0g
  • DMSO: 2.5ml
  • MgCl2[1M]: 0.25ml
  • MgSO4[1M]: 0.5ml
  • 100% glycerol: 7.5ml

Store all competent cells at -80 deg C for future use.

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