1. Grow cells to an OD660 ~ 1 UA/ml.
2. Spin cells at 2500g for 1 min. Wash pellets in an equal volume ofice-cold water.
3. Wash in ½ volume cold water (for 40 ml culture, 20 ml water).
4. Wash in 1/25 volume cold water (for 40 ml culture, 1.6 ml water).
5. Treat cells with 25 mM DTT for 10 minutes at room temperature.
6. Wash cells with 1 M cold sorbitol.
7. Resuspend cells in 1/200 original volume in cold 1 M sorbitol (for 40 ml culture, 200 μl 1 M sorbitol).
8. Mix 50 μl of cell suspension with no more than 5 μl DNA (in low ionic strength buffer).
9. Immediately tap cell/DNA suspension to the bottom of a 0.2 cm cuvette, pulse at 1.5 kV, 200 Æ, 25 μFaradays (pulse time must be about 5 ms).
10. Immediately add 1 ml YPD/1M sorbitol and allow recovering at permissive temperature for one hour.
11. Spin down cells and resuspend in 1 M sorbitol.
12. Plate on selective medium (medium should contain 1 M sorbitol).
Note that 10 ml is the minimum culture volume and that cells can be stored for a few days if they are not DTT treated.