Dominik Haudenschild, October 14, 1997

2) Heat inactivate ligation mix 10-15 minutes at 68°C

3) Put 30-40µl of cells + 1µl of ligation mix into sterile tube, mix gently

4) Put electroporation chamber (0.1cm) into ice

5) Put cells + DNA into electroporation chamber

6) Adjust electroporator & power supply settings

Power Supply: 1500 Volts (limiting), 25mA, 25W

Electroporator: 50µF, 150W , Disarmed, Charge

7) Electroporate

a) Place Chamber into electroporator & close lid

b) Charge until mA reading is steady (~5.9 to 6.2)

c) Turn switch to "Armed", red light goes on

d) Push switch to "Pulse"

e) Return switches to "Disarmed" and "Charge"

f) Place Chamber on Ice for 1-2 minutes

8) Add cells to 1ml SOC, 37°C, no antibiotics

9) Shake in 37° incubator 1 hour

10) Plate onto 3 150mm LB-agar plates with appropriate

antibiotics. Plate 1: 100µl of cells

Plate 2: 250µl of cells

Plate 3: Rest of cells

11) Incubate overnight 37° incubator then store @ 4°C no more

than 4 weeks before re-plating