Effects of Lipopolysac-charide on Transfection Efficiency in Eukaryotic Cells

摘要

The suitability of different purification methods for preparation of plasmid DNA for transfection into eukaryotic cells was systematically investigated. The reporter plasmid, pRSVcat, was prepared using several methods, and residual impurities in thepreparations were quantitated, Transfection with these preparations was performed with several cell lines (HeLa, Huh7, COS7 and LMH) and two transfection methods: lipo-some-mediated and calcium phosphate transfection. Transfection efficiencies were determined by measuring chloramphenicol acetyltransferase expression. Higher transfection efficiencies were obtained with plasmid preparations of higher purity (those prepared by union-exchange chromatogra-phy or two rounds of CsCl-gradient cen-trifugation) than with preparations of lower purity (those prepared using a silica-based DNA adsorption method or a single round of CsCl cenlrifugation). The results also demonstrated specifically that increasing concentrations of lipopolysaccharides in plasmid preparations directly correlate with decreasing transfection efficiencies.