Inoculate 1 liter of LB medium with 5 ml of an overnight culture of bacteria and grow at 37. When the OD600 reaches 0.5, heat shock the culture for exactly 15 min at 42°C (with agitation) and chill the culture in ice slurry for 10 min. For now on, all procedures are made at 4°C. Harvest cells at 10.000 rpm for 3 min. Resuspend in 100 ml of ice-cold 10% glycerol and centrifuge again. Repeat this step once again. Resuspend bacteria to a final volume of 2 ml in ice cold GYT (10% glycerol, 0.125% yeast extract and 0.25% tryptone). … Continue reading Electrotransformation of E. coli