Difference between revisions of "Team:Birkbeck/Chemical Transformation"
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Latest revision as of 17:28, 18 September 2015
Chemical Transformation
See previous protocol: Chemical Competent E. coli Cell Protocol for details on making competent E. coli culls.
Chemical Transformation protocol.
- 1. Thaw out the 50 µL cell suspension aliquot on ice.
- 2. Add 100pg-1µg of plasmid DNA to the cell suspension (approximately 1-5 µL of plasmid solution).
- 3. Mix plasmid and cells thoroughly and incubate on ice for 30 minutes.
- 4. Induce heat shock by incubating cells at 42oC for 30 seconds.
- 5. Incubate on ice for 5 minutes.
- 6. Add 450 µL of LB and incubate at 37oC for 1 hour.
- 7. Plate out 50 µL of straight transformant cells & also carry out a 10-fold dilution (plate out 50µL). Note that the plate should contain the appropriate antibiotic at the MBC in order to select for transformants which contain the desired plasmid.
- 8. Incubate overnnight at 37oC in a static incubator.