Experimental strategies in efficient transfection of mammalian cells. Calcium phosphate and DEAE-dextran.

Abstract

Transfection of foreign DNA into mammalian cells is one of the most common procedures performed by investigators next to cell culture itself Whether the goal IS to express mammalian or viral genes transiently or by the generation of stable cells lines, researchers have a variety of transfection methods to choose from Calcium phosphate coprecipitation and DEAE-dextran-mediated transfection are the oldest, and with many investigators, the most trusted of these methods. Paradoxically, the exact mechanism of DNA uptake for each of these methods remains unknown DEAE-dextran mediated DNA transfection is a favored approach for transient expression experiments The method has evolved somewhat since its original description (1-^). In general this method is highly reproducible, more so than calcium phosphate transfection, and can be performed, with some optimization, on a wide variety of cell types. The efficiency of transfection can be as high as 80% with some cell types (5). The principle drawback to DEAE-mediated DNA transfection is the poor efficiency of forming stably integrated expressing cell lines. Thus, DEAE-dextran transfection can only be used for transient expression assays. Other limitations include cellular toxicity, necessity for a DMSO (dimethyl sulfoxide) shock to improve efficiency, and the additional constraint that only plasmid DNA gives good transfection efficiencies. In addition, cotransfection efficiencies are not as good as those achieved with the calcium phosphate method. Notwithstanding these limitations, DEAEdextran-mediated DNA transfection is a widely used procedure that can be made