I typically use this protocol (https://www.neb.com/protocols/1/01/01/electroporation-protocol-c2986
) from New England Bio labs. We run it as follows.
1. Bring culture tubes with a round bottom (we use VWR #60818-667) to room temperature. Warm the SOC recovery medium to 37°C in a water bath. Warm your selection plates to 37°C for 1 hour.
2. Put 1 mm electroporation cuvettes (1mm) and microcentrifuge tubes for cells and DNA in ice bucket with ice.
3. For a transformation positive control, dilute the control pUC19 with sterile water by 1:5 to 10 pg/μl final concentration.
4. Thaw electrocompetent cells (we use the New England Bio labs turbo ones) on ice for approx 10 minutes and flick the tubes gently to mix the cells. Transfer 25 μl of the cells to one of the chilled microcentrifuge tubes from step 2 . Add 1 μl of your DNA solution to the chilled tube with the cells.
5. Transfer the mix of cells and DNA from the microfuge tube into a chilled electroporation cuvette, ensuring that there are no bubbles and the cells deposit on the bottom of the cuvette. Electroporate using the using the following settings: 2.1 kV, 100 Ω, and 25 μF. Our lab usually uses a time constant of ~2.6 ms.
6. Immediately after electroporation, pipette 975 µl of 37°C SOC (from step 1) to the cuvette, gently mixing the solution up and down twice, then transfer the contents of the cuvette to the culture tubes from step 1.
7. Shake the culture tubes vigorously at 250 rpm or rotate continuously for 1 hour at 37°C.
8. Dilute the cells to desired concentration and spread 100-200 μl cells onto a pre-warmed selection plate (from step 1).
9. Incubate selection plates 8 -12 hours at 37°C.