Optimization of electroporation for transfection of mammalian cell lines.
Electroporation can be a highly efficient method for introducing DNA molecules into cultured cells for transient expression of genes or for permanent genetic modification. However, effective transformation by electroporation requires careful optimization of electric field strength and pulse characteristics. We have used the transient expression of the firefly luciferase gene as a rapid and sensitive indicator of gene expression to describe the effects on transfection efficiency of altering electroporation field strength and shape. Using the luciferase assay, we investigated the correlation of cell viability with optimal transfection efficiency and determined the optimal parameters for a number of phenotypically distinct mammalian cell lines derived from the nervous and immune systems. The efficiency of electroporation under optimal conditions was compared with that obtained using DEAE-dextran or calcium phosphate-mediated transformation. Transfection by electroporation using square wave pulses, as opposed to exponentially decaying pulses, was found to be significantly increased by repetitive pulses. These methods improve the ability to obtain high efficiency gene transfer into many mammalian cell types.
Andreason
G L
GL
Molecular Genetics Laboratory, Salk Institute, La Jolla, California 92138-9126.
Evans
G A
GA
D000818:Animals; D002130:Calcium Phosphates; D003637:DEAE-Dextran; D004574:Electromagnetic Fields; D008322:Mammals; D008722:Methods; D051379:Mice; D051381:Rats; D013696:Temperature; D014162:Transfection; D014407:Tumor Cells, Cultured
D002130:Calcium Phosphates; C485828:alpha-tricalcium phosphate; C485830:tetracalcium phosphate; C485838:calcium phosphate, monobasic, anhydrous; D003637:DEAE-Dextran; C020243:calcium phosphate; C485829:calcium phosphate, dibasic, anhydrous