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TOP10 chemically competent cells: Difference between revisions
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TOP10 chemically competent cells (view source)
Revision as of 22:36, 8 November 2006
, 8 November 2006→Preparing competent cells
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===Preparing competent cells=== | ===Preparing competent cells=== | ||
* Inoculate 250 ml of [[SOB]] medium with 1 ml vial of seed stock and grow at 20 C to an OD of 0. | * Inoculate 250 ml of [[SOB]] medium with 1 ml vial of seed stock and grow at 20 C to an OD of 0.3 | ||
** This takes approximately 16 hours. Controlling the temperature makes this a more reproducible process, but is not essential. Room temperature will work. You can adjust this temperature somewhat to fit your schedule | ** This takes approximately 16 hours. Controlling the temperature makes this a more reproducible process, but is not essential. Room temperature will work. You can adjust this temperature somewhat to fit your schedule | ||
* Centrifuge at 3000g / 4C for 10 minutes in a flat bottom centrifuge bottle. | * Centrifuge at 3000g / 4C for 10 minutes in a flat bottom centrifuge bottle. | ||
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* Store at -80C indefinitely. | * Store at -80C indefinitely. | ||
** Flash freezing does not appear to be necessary | ** Flash freezing does not appear to be necessary | ||
===CCMB80 buffer=== | ===CCMB80 buffer=== | ||
