Electroporation

Making Competent Cells

Materials

• N samples of DNA to be transformed, 1 positive control sample
• N+1 electroporation cuvettes
• N+1 50μL aliquots of EC cells (we used DH5α)
• N+1 mL SOC (Super-optimal broth + 10mM glucose)
• N+1 LB-agar plates of appropriate resistances

Procedure

1. Chill electroporation cuvettes and thawed EC cells on ice. Do not let EC cells warm to above ice-cold.
2. Add 1μL of ligation product to the 50μL aliquot. Don't forget an aliquot for positive control.
3. Transfer DNA/cell mixture to a cuvette and pulse at 2.5V. Rescue the cells immediately by adding 0.75mL warm SOC to the cuvette.
   1. Ensure that the time constant is above 3.0 for each.
4. Incubate for 1 hour at 37°C.
5. Plate the entire mixture on an LB-agar plate with the appropriate antibiotic resistance, grow overnight.