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Unformatted text preview: March 25, 2009 Transformations with chemically competent cells Chemically competent cells can be made via many procedures. We use the Inoue protocol for supercompetent bugs as documented in Maniatis/Sambrook. Also, transformation protocol depends upon the strain of bacteria to some extent. The most common ones in our lab are XL1- Blue mrf’ (Stratagene) and occasionally DH5 α . We have stored our chemically competent cells in batches of 200-300 μ l in microfuge tubes at –80ºC. Use 100 μ l (you can get away with 50 μ l) per transformation. XL1-BLUE chemically competent cells marked with blue “C” . XL1-BLUE electrocompetent cells marked with blue “E” . DH5 α chemically competent cells marked with red “C” . DH5 α electrocompetent cells marked with red “E” . General procedure: First, set a water bath to 42ºC!!!!!!! 1. Have DNA from ligation or from plasmid prep ready . For plasmids, amount is not critical, but you can use on the order of 1-10 ng and get plenty of colonies. critical, but you can use on the order of 1-10 ng and get plenty of colonies....
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This note was uploaded on 11/25/2009 for the course BI 379 taught by Professor Kavaler during the Spring '09 term at Colby.
- Spring '09