Some of the simple techniques useful for students like me.
- Overnight 5ml subculture at 37 ℃ for 18hr.
- Inoculate 100µl subculture into 100 ml LB broth.
- Culture at 37 ℃, 2½ hr.
- Centrifuge at 4000 rpm, 10mins at 4°C (50ml +50ml).
- Re-suspend cell pellet in 5ml 0.1 M cacl2 for each tube.
- Keep on ice for 5mins.
- Finally re-suspend cell pellet in 3 ml 0.1 m Cacl½ (1.5ml for each )
- Preserve at -70 ℃ by adding 600µl of 50% glycerol ( final 10% gly) and aliquot into 1.5 ml tubes containing 200µl each.