Berglund:CaCl2 cells

Preparation of CaCl2 (heat-shock) competent cells

row 500ml-1L to an OD600 between 0.4 ad 0.6 Place cells on ice for 15 minutes spin down cells 4000rcf for 10 minutes resuspend gently in half the pellt volumes of ice cold 50mM CaCl2 Let sit on ice for 30 minutes spin down cells 4000rcf for 10 minutes resuspend the cells gently in 1/20 of the pellet volume in ice cold 50mM CaCl2 + 10% glycerol incubate at least one hour on ice make 300µl aliquots (use 100µl per transformation) quice freeze in a dry ice/ethanol bath or liquid nitrogen store at -80. Transformation protocol add 1µl (100ng) plasmid and heat shock for 45 seconds at 42 degrees in a water bath