Transfection of the ChloramphenicolAcetyltransferase Gene into Eukaryotic Cells Using Diethyl-Aminoethyl (DEAE)-Dextran

The study of gene regulation in eukaryotic cells involves a practical requirement for two distinct techniques: first, a transfection system, or more simply, a way of getting DNA into a cell; and second, a reporter system, which, as the name suggests, is a means of finding out what the transfected DNA does from its new location inside the cell. These two requirements are amply met by transfecting cells, in the presence of the polycation diethyl aminoethyl-dextran (DEAE-dextran), with a plasmid vector that has regulatory sequences adjacent to the chloramphenicol acetyl transferase (CAT) gene. This type of gene transfer is often referred to as a transient expression system. Production of the CAT enzyme peaks at around 40–48 h and thereafter the level falls as the plasmid DNA is diluted out in a growing population of cells. The general strategy of subcloning putative regulatory regions followed by transfection and quantification of CAT activity is outlined in a flow diagram (Fig. 1 )

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