Inclusion of high molecular weight dextran in calcium phosphate-mediated transfection significantly improves gene transfer efficiency.
Calcium phosphate-based mammalian cell transfection is a widely used gene transfer technology. To facilitate the efficiency of this gene transfer method, several polysaccharide compounds were tested and evaluated for their effectiveness in enhancing DNA transfection. Using a HIV-1-derived lentivirus vector plasmid as a gene transfer indicator, we demonstrated that the addition of high molecular weight dextran-500 at 0.6-1.2% in the 2x Hepes buffered saline (HBS) increased transfection efficiency by over 50% (as reflected by the number of GFP-positive cells) and increased the titer of resulting lentivirus vector particles even more (up to 4-fold). This enhancement of transfection efficiency was further increased when higher molecular weight dextran formulations were used in place of dextran-500, and also when dextran was used in combination with polybrene, another polycationic chemical compound. Examination of transfected cells showed that dextran had no apparent adverse effect on cell viability and growth. Our data represent the first report showing that dextran can be used to enhance calcium phosphate-mediated gene transfer; this may be useful in applications for the generation of high-titer virus vector stocks using transient transfection technology.
Wu
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C
Department of Public Health Sciences, University of Hawaii at Manoa, Honolulu, Hawaii 96822, USA.
Lu
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Y
D002130:Calcium Phosphates / Q000494:pharmacology*; D002478:Cells, Cultured; D004247:DNA / Q000494:pharmacology; D003911:Dextrans / Q000737:chemistry / Q000494:pharmacology*; D004526:Efficiency; D006801:Humans; D008970:Molecular Weight; D009994:Osmolar Concentration; D010957:Plasmids / Q000235:genetics; D011073:Polyamines / Q000494:pharmacology; D000071228:Polyelectrolytes; D014162:Transfection / Q000379:methods*
D002130:Calcium Phosphates; D004247:DNA; D003911:Dextrans; D011073:Polyamines; D000071228:Polyelectrolytes; C020243:calcium phosphate