Transfection of Human Breast Epithelial Cells with Foreign Dna Using Different Transfecting Techniques

Abstract

The introduction of DNA into eukaryotic cells is a powerful technique for the study of gene regulation. This requires both a technique by which genes can efficiently be introduced into cells and a recipient cell representative of the tissue under study. We have utilized a spontaneously immortalized human breast epithelial cell line, CMF-10, which is phenotypically normal for introducing a neomycin-resistant gene contained in the plasmid Homer 6 (pHo6). Three different transfection methods were tested: calcium phosphate, with and without glycerol shock, using as control NIH/3T3 cells, and strontium phosphate and electroporation. In MCF-10 cells the efficiency of calcium phosphate mediated DNA transfection, which was measured as the number of colonies growing in neomycin-containing medium, was 21 and 140 fold higher than in MCF-10 cells transfected by electroporation and strontium phosphate methods, respectively. Glycerol shock enhanced three fold transfection efficiencies. NIH/3T3 transfected cells by calcium phosphate method showed a transfection efficiency similar to that of MCF-10 cells, however, glycerol shock did not improve the efficiency. These studies revealed that calcium phosphate combined with glycerol shock is the most efficient technique for transfection of genomic DNA into human breast epithelial cells.