"
Team:Newcastle/E. coli Competence
From 2010.igem.org
Revision as of 14:42, 29 July 2010 by Shethharsh08 (Talk | contribs)
| |||||||||||||
| |||||||||||||
Competent cells preparation
Materials required
- Conical flask
- 300ml of LB broth
- 1/40 volume of an overnight culture
Procedures
- Inoculate 300 ml of LB broth in a conical flask and inoculate with 1/20 volume of an overnight culture of the desired strain.
- Grow the cell at 37°C in an incubator(with a shaking platform so as to mix the media equally amongst the cells).
- Chill cells by placing the flask on ice and harvest by centrifugation at 4°C for 10 minutes.
- Resuspend the pellet in 100 ml of ice cold TFB1 and gently shake the tubes whilst placed on ice.
- Repeat the the above mentioned step and carefully resuspend pellet in 20 ml ice cold TFB2.
- Aliquot 200 µl volumes of the cell suspension into cold sterile microfuge (1.5 ml) tubes and flash freeze in dry-ice
- Store at -80°C
   
|
    
|
