Use of a quaternary ammonium detergent in liposome mediated DNA transfection of mouse L-cells.
Sonicated liposomes composed of dioleoylphosphatidylethanolamine (DOPE) and a quaternary ammonium detergent (dodecyl-, tetradecyl-, or cetyl-trimethylammonium bromide) mediates functional transfer of pSV2 CAT plasmid DNA to mouse L929 fibroblasts. Successful transfection was determined by assaying for chloramphenicol acetyltransferase activity in cell lysates collected 40 h after exposure to the lipid-DNA complexes. Liposomes prepared with the quaternary ammonium detergents were less toxic than the free detergents at the same concentrations and were more efficient in their delivery of the plasmid DNA to the cells. Analysis of the three detergents in combination with the lipid showed that cetyltrimethylammonium bromide was least toxic to the cells. This detergent, at a minimal concentration of 20 mol% in DOPE, allowed for stable liposome preparations and efficient transfection. Optimal efficiency of transfection occurred with 30 micrograms of DNA. Further increases in the DNA concentration caused a decrease in the transfection efficiency, perhaps due to charge repulsions between the liposomes now saturated with negatively charged DNA and the negatively charged cell surface. The transfection activity of the liposome was limited by its cytotoxicity at high liposome concentrations. These results are compared with that of the Lipofectin, another positively charged liposome preparation which is commercially available. Although the overall transfection activity of the liposome containing the quaternary ammonium detergent is somewhat lower than that of the Lipofectin, it may serve as an inexpensive and convenient alternative.
Pinnaduwage
P
P
Department of Biochemistry, University of Tennessee, Knoxville 37996-0840.
Schmitt
L
L
Huang
L
L
D000818:Animals; D002478:Cells, Cultured; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D004247:DNA / Q000235:genetics*; D003902:Detergents; D015870:Gene Expression; D007739:L Cells / Q000201:enzymology; D008081:Liposomes / Q000378:metabolism*; D051379:Mice; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010957:Plasmids; D000644:Quaternary Ammonium Compounds / Q000494:pharmacology*; D013501:Surface-Active Agents; D014162:Transfection
D003902:Detergents; D008081:Liposomes; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D013501:Surface-Active Agents; C020888:1,2-dielaidoylphosphatidylethanolamine; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase