ADENOVIRUS-MEDIATED TRANSFECTION OF CULTURED-CELLS

We describe here a simple and efficient transfection method for transi ent expression of cloned genes in cell lines and primary cultured cell s. The method involves the use of DEAE-dextran to target DNA to the ce llular endocytotic pathway and the use of a human adenovirus to ensure efficient lysis of endosomal vesicles. The procedure allows effective delivery of DNA into the cytoplasm and, therefore, results in a highe r fraction of cells expressing exogenous proteins. Using this method w e routinely obtain 60%-90% of COS cells or Chinese hamster ovary cells expressing beta-galactosidase, as determined by in situ staining with 5-bromo-4-chloro-3-indolyl-beta-D-galactoside (X-gal). We have also o btained much improved levels of expression in cells that are difficult or impossible to use in transient expression assays, such as rat-1 fi broblasts or primary osteoblast cultures. We successfully used the met hod to express heteromeric proteins that require subunit assembly for proper function. The method also proved effective to express functions in which the exogenous protein needs to couple to the endogenous cell ular machinery Thus, this transient transfection method should prove v aluable for many functional studies in a broad variety of cell lines a nd primary cultures.