Protocols

Midi-prep (fast but ± clean)

  1. Transfer 25 mL of bacterial culture in a 50 mL falcon
  2. Centrifuge 10 minutes at 4ºC
  3. Discard supernatant
  4. Resuspend pellet in 2 mL of solution I
  5. Add 2 mL of solution II (mini-prep) DO NOT VORTEX
  6. Add 2 mL of chloroforme
  7. Wait 1 minute at room temperature
  8. Add 1.5 mL of solution III
  9. Centrifuge 10 minutes at 4ºC
  10. Transfer surnatant in a new falcon (avoid transfering cell debris)
  11. Prepare a mix phenol/chloroforme 1:1
  12. Add an equal volume of phenol/chloroforme to supernatant
  13. Centrifuge 10 minutes at 4ºC
  14. Transfer supernatant
  15. Add 2 volume of EtOH 100%
  16. Add 1/10 (final volume of precedent step) of sodium acetate 3M pH 5.2
  17. Centrifuge 10 minutes at 4ºC
  18. Wash with 10 mL EtOH 70%
  19. Centrifuge 10 minutes at 4ºC and keep pellet
  20. Air-dry pellet completely
  21. Resuspend in H2O with RNAse (40ug/mL)
  22. Dissolve pellet 5 minutes at 65oC or 45 minutes at 37ºC
Modified from a Midi-prep protocole of Jean-Claude Labbé.