Seed cells at appropriate density (5x104/well for 3T3, MPAC, BHK; 5x105/well for aTC1.6, bTC3) one to two days before transfection.
Combine total DNA at 1ug /well with 600ul/well serum-free medium in an eppendorf tube.
Add 1.5ul/well Transfast (Promega) to the DNA/medium mixture. Vortex for 10 seconds.
Incubate at RT for 10 to 15 min.
During the above incubation, aspirate medium from cells and add PBS; remove PBS just before adding DNA in step 6.
Add 600ul DNA mixture from step 3 to the appropriate well.
Incubate at 37 oC incubator for 1 hr.
Add 4ml regular growth medium to each well.
after 24 hours, change to fresh medium.
After 48 hrs, harvest cells for functional assay.
