Making electrocompetent cells:
1. Grow cells to mid-exponential growth phase at an O.D.650of 0.5. 2. Harvest by centrifugation at 3,000 x g for 10 min at 4 °C. 3. Wash twice in 15% glycerine at 4 °C. 4. Resuspend in 1/20 volume of 15% glycerine and keep at 4 °C.
Electroporation of cells:
1. Add 3 µl (300 µg/ml) plasmid DNA to 125 µl of electrocompetent cells. Homogenize by gently mixing with pipette several times. Transfer mixture into a prechilled cuvette. 2. Wipe moisture from the cuvette and insert the cuvette into the device. 3. Electroporation:
Expected results:
Transformation efficiency up to 1 x 107 transformants/µg of DNA.
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