Team:Wageningen UR/Protocol/StartupCCMV
From 2012.igem.org
Revision as of 10:38, 10 September 2012 by MarkvanVeen (Talk | contribs)
Growing Culture
Reagents & Materials
For a 100 ml batch:
reagents:
Procedure
- Prepare a Erlenmeyer flask with 0.1 L of LB-medium and keep it at 37°C
- Pick E.coli BL21 from a plate and grow them over night in a 10 mL LB culture containing 50mg/L Kanamycin at 37°C
- Inoculate the large flask with 1 mL of the overnight culture and grow at 37°C for 2.5h until the OD600 reaches approx. 0.6
- Induce with 1.25 mM IPTG (0.5 mL of a 250 mM stock)
- Incubate at 37°C for 4 h
- Centrifuge the cells in 50ml greiner tubes at 4700 rpm for 18 minutes
- Decant the supernatant and resuspend the pellets in ± 20 mL mQ water each
- Centrifuge again at 4700 rpm for 18 minutes
- Decant the supernatant and centrifuge for another minute
- Pipette any liquid to clear the tube of supernatant
- (possible to freeze the cells to -20°C at this point)
Continue further for CCMV formation