Introduction of DNA into 3T3-L1 Adipocytes by Electroporation

Abstract

After cloning a gene of interest, many researchers wish to analyze its characteristics by overexpression analysis or by introduction of mutated forms of the gene of interest into various cell types. In the analysis of insulin-stimulated glucose transport, the most appropriate cell systems are striated muscle and adipocytes (1). However, the introduction of DNA or genes of interest into these insulin-responsive tissues by standard transfection protocols such as calcium phosphate, DEAE-dextran, and liposome-mediated transfection are very inefficient. Furthermore, although transgenes can be expressed in muscle using adenovirus infection systems, this is difficult to accomplish in adipocytes and is substantially more labor intensive. The production of recombinant adenoviruses to use in infection of insulin-responsive tissues can take several months and requires very high titers of adenovirus. Therefore, we have recently established electroporation conditions that consistently provide at least 50% transfection efficiency for cultured differentiated 3T3-L1 adipocytes (2). Although the electroporation is not 100% efficient, it provides an easy and fast method to introduce DNA into adipocytes. Using 600 μg of CMV-LacZ plasmid DNA, we consistently obtain an electroporation efficiency of 50–80% (see Fig. 1).