IGEM:PennState/2008/PreparingElectrocompetentCells

Electrocompetent Cell Prep

Starter Culture

• Inoculate desired cell strain (DH5a) into 3 mL of LB and shake at 37C

Prep

1. OD the starter culture, use to calculate the dilution into 30 mL of LB for desired growth time (~30 min doubling time).
2. Shake in falcon tubes at 37C until OD of 0.5-0.6
3. Put cells in ice water bath for 30 min
4. Wash Cells
   1. Spin down cells in rotor (max rpm)at 4C for 10 min
   2. Pour off supernate and re suspend cells in 30mL of cold water
   3. Repeat wash 2 times
   4. After final spin down, pour off the supernate and re suspend cells in ~300uL water
5. Use 70uL allloquats for transformations