Trinh:Chemically Competent Cells
BE SURE TO PERFORM THIS IN A STERILE ENVIRONMENT. READ AND PREPARE BEFORE PERFORMING.
- Inoculate 10 ml of SOB medium with seed stock and grow at 37°C to an OD600nm of 0.5
- Place inoculum on ice for 30 minutes
- Shake frequently to maintain homogeneous solution
- Centrifuge at 4000rpm at 4°C for 10 minutes.
- Discard supernatant by pouring out slowly and pipeting remaining supernatant.
- Gently resuspend in 10 ml of ice cold CCMB80 buffer.
- Incubate on ice 20 minutes
- Centrifuge again at 4°C and discard supernatant as described above.
- Resuspend in 10 ml of ice cold CCMB80 buffer to an OD of ~3.0
- Aliquot into chilled screw top or 1.5 mL graduated tubes
- Store at -80°C.