Purifying Chromosomal DNA from E. coli

JOEL'S PROTOCOL

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1. Pellet 1 ml of O/N.
2. Wash with TNE.† (Use the same volume of TNE as O/N pelleted in step#1)
3. Spin down and decant supernatant.
4. Resuspend in 135 uL of TNE + 2% Triton X-100 per ml of O/N culture.
5. Add 0.15 mg of lysosyme per ml of O/N culture.
6. Incubate at 37 deg C †for 30 minutes.
7. Add 0.3 mg of proteinase K per ml of O/N culture.
8. Incubate at 65 deg. C for 2hrs.

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*Use 3 uL for a Southern Blot and 2 uL for PCR reactions.

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TNE

10mM Tris-HCl pH 8.0

10mM NaCl

10mM EDTA pH 8.0