ELECTROPORATION-MEDIATED TRANSFECTION OF MAMMALIAN-CELLS WITH CRUDE PLASMID DNA PREPARATIONS

We designed a simple and reproducible electroporation-mediated transfe ction procedure with which to screen mammalian expression vector-const ructed cDNA libraries. Using a specific chamber composed of five paral lel electrodes, the recipient cells can be electroporated separately w ith 40 plasmid DNA preparations in a single experiment. Over 300 crude plasmids prepared From E. coil (DH-5) carrying a pcD2neo-vector-deriv ed cDNA library were tested. The efficiency of stable transfection by electroporation with crude plasmid DNA preparations was 10-times highe r than with the CsCl-purified plasmid DNA. When the crude plasmids wer e digested with RNase, the efficiency of stable transfection markedly decreased, indicating that the contaminating bacterial RNA in the crud e plasmid preparations has a strong carrier effect during the electrop oration. Even when salmon sperm DNA or genomic DNA from the recipient cells was used as the carrier of the purified plasmid, the efficiency was not higher than that using the crude preparations. This procedure is useful not only for screening a number of cDNAs but also for routin ely introducing biologically active foreign genes into cultured mammal ian cells.