Control by pulse parameters of electric field-mediated gene transfer in mammalian cells

Abstract

Electric field-mediated gene transfer in mammalian cells (electrotransformation) depends on the pulsing conditions (field intensity, pulse duration, number of pulses). The effect of these parameters was systematically investigated using the transient expression of the chloramphenicol acetyltransferase and the beta-galactosidase activities in Chinese hamster ovary cells. Pulsing conditions inducing reversible permeabilization 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 state 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.