Template:SBB-Protocols Micro1
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Transformation by heat-shock
Competent cells are stored as 200uL aliquots in the -80 freezer as a communal stock.
- Thaw a 200 uL aliquot of cells on ice
- Put your ligation mixture on ice
- Add 50 uL of water to the mixture
- Add 30 uL of KCM salts to the mixture
- Add 75 uL of the cell cocktail to the ligation, pipette up and down gently to mix
- Let sit on ice for 5 min
- Heat shock for 2 min at 42
- Put back on ice for 1 min
- For ampicillin selection, you can plate immediately, otherwise:
- Add 100uL of LB, let shake in the 37 degree incubator for 40 min
- Plate on selective antibiotics, let incubate overnight