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Miniprep of Bacterial Genomic DNA

  1. Grow bacterial strain to saturation
  2. Spin 1.5 mL for 2 min in microcentrifuge
  3. Resuspend in 567 ul TE buffere, 30ul of 20 mg/ml proteinase K. Mix and incubate 1 hr at 37C
  4. Add 100ul of 5M NaCl. Mix thoroughly
  5. Add 80 ul of CTAB/NaCl solution. Mix. Incubate 10 min at 65C.
    • CTAB/NaCl solution (0.7M NaCl, 10% CTAB): Dissolve 4.1 g NaCl in 80 ml water. Slowly add 10 g CTAB. Stir with heat to dissolve. Bring volume to 100 ml.
  6. Extract with an equal volume of chloroform/isoamyl alcohol. Spin 5 min in microcentrifuge.
  7. Transfer aqueous phase to a fresh tube. Precipitate DNA with .6 vol isopropanol. Wash precipitate with 70% ethanol. Remove supernatant and briefly dry pellet in lyophilizer.
  8. Resuspend pellet in 100ul TE buffer.