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Bacterial Glycerol Stock.html
Bacterial Glycerol Stock

Materials

  1. Agar plate with isolated colonies
  2. LB media
  3. Glycerol
  4. Sterile flasks with ventilating caps or 50 ml tubes
  5. Screw-cap freezer vials
  6. Shaker incubator
  7. 5mL serological pipettes
  8. Micropipette and sterile tips
  9. Ethanol-dry ice bath
  10. -80oC freezer

Procedure

  1. Fill 5mL media each into the flask/tube, aseptically.
  2. Add antibiotics if needed.
  3. Pick a single colony from agar plate and disperse into the media.
  4. Grow overnight culture of bacteria at 37oC and 300rpm.
  5. Add sterile glycerol (autoclaved) to 15% v/v final.
  6. Vortex vigoriously and dispense 1ml aloiquots in screw-cap freezer vials.
  7. Quick freeze on Ethanol-dry ice bath.
  8. Transfer to -80oC freezer for long term storage.