"
Team:SDU-Denmark/Protocols/Competent-cells
From 2009.igem.org
Protocol for making competent cells (E.coli)
(Cells are kept on ice at all times!! If the cells temperature rises above ~5º C they'll lose their competency!)
- 600 µl overnight (ON) Top10 E. coli culture is added to 60 ml Luria-Bertani (LB) medium.
- Grows at 37º C while being shaken until the optic density (OD550) is 0,2.
- Cool cells on ice.
- Harvest the cells in screwcap tubes (4 × 10 ml).
- Pour away the supernatant and keep the pellet on ice.
- Wash the cells with 10 ml cold 50mM CaCl2.
- The cells are being distributed in eppendorf tubes of 200 µl.
- Add 41.7 µl 87% glycerol and mix well.
- Store at -80º C.
