Method
- Pick a colony into 2 ml L-Broth containing the appropriate
antibiotics and incubate at 37°C with shaking overnight.
- Spin down 1 ml culture in Eppendorf tube for 5 minutes (at room
temperature).
- Resuspend pellet in 45 µl of STET buffer add 5 µl lysozyme (10
mg/ml).
- Put in boiling water for 2 minutes.
- Spin down for 10 minutes (at room temperature) in Eppendorf
centrifuge.
- Transfer supernatant to a fresh tube, add equal volume of
isopropanol and mix.
- Spin down at 12000 RPM, 4°C for 30 minutes. Pour off supernatant.
- Air dry the tube and resuspend in the appropriate buffer for
digestion with restriction enzymes.
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