Transformation of Cells
Objective
Transformation of BL21(DE3) cells with plasmid DNA (T39C L72C Mb) for ampicilin resistance.
Procedure
- Thaw on vial of cells on ice
- Add 5-10ng of DNA, in a volume of 1-5uL to the cells and mix by tapping gently. Do not mix cells by pipetting.
- Incubate the vial on ice for 30 minutes
- Heat shock the cells by incubating the vials for exactly 30 seconds in the 42 degree Celsius water bath. Do not mix or shake.
- Remove the vial from the 42 degree Celsius water bath and quickly place on ice.
- Add 250uL of pre-warmed SOC medium to the vial. (SOC is a rich medium; use proper sterile technique to avoid contamination.)
- Secure the vial in a micro centrifuge rack with tape. Place the rack in a shaking incubator, and shake the vial at 37 degree Celsius for 1 hour at 225 rpm.
- Plate volume of cells onto LB plates containing appropriate antibiotic for plasmid selection.
- Invert the plates and incubate at 37 degrees Celsius overnight
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