Materials

1. Agar plate with seperated colonies
2. LB liquid media
3. Sterile tubes or flasks with ventilaing caps
4. Shaker incubator
5. Pipette and sterile tips

Procedure

1. Put 5ml media into the tube, aseptically.
2. Add antibiotic if needed.
3. Pick a single colony from agar plate and disperse into the media.
4. Grow overnight culture of bacteria at 37^oC 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 dry ice.
8. Transfer to -80C freezer for long term storage.