CONTROL BY PULSE PARAMETERS OF ELECTRIC FIELD-MEDIATED GENE-TRANSFER IN MAMMALIAN-CELLS

Electric field-mediated gene transfer in mammalian cells (electrotrans formation) depends on the pulsing conditions (field intensity, pulse d uration, number of pulses). The effect of these parameters was systema tically investigated using the transient expression of the chloramphen ycol acetyltransferase and the beta-galactosidase activities in Chines e hamster ovary cells. Pulsing conditions inducing reversible permeabi lization of the cell plasma membrane are not sufficient to induce gene transfer. The plasmid must be present during the electric pulse if it is to be transferred across the membrane into the cytoplasm. Only the localized part of the cell membrane brought to the permeabilized stat e by the external field is competent. Pulse duration plays a key role in the magnitude of the transfer. The field induces a complex reaction between the membrane and the plasmid that is accumulated at the cell interface by electrophoretic forces. This leads to an insertion of the plasmid, which can then cross the membrane.