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Use of plasmid DNA purified by PEG precipitation for in vivo transfection in mice

Abstract:

Background. To demonstrate that plasmids obtained by PEG 6000 precipitation method can be used for in vivo transfections. Materials and methods. The mammalian expression vector for beta-galactosidase (β-gal) was purified following the PEG 6000 precipitation protocol, purified DNA was assayed in transfection experiments in COS-7 cell line by DEAE-Dextran procedure. Two or three doses of 100 μg of plasmids were injected into murine skeletal muscle and the expression of the β-gal gene was examined. In addition we evaluated seroconversion by ELISA and these results were confirmed in a competition assay using soluble β-gal. Results. We obtained an average purification yields of 14.31±1.5 mg/L of culture; with 90% to the molecules in the supercoiled form and 260/280 ration between 1.8 and 1.9. Transfection efficiency in COS-7 cells, ranged between 25-30%. The 85% of the animals have expressed β-gal in muscle and seroconversion has been demonstrated in 28% of the injected mice. Conclusions. We have demonstrated that plasmids obtained by PEG 6000 precipitation method can be successfully used for in vivo transfection. This technique is less expensive and laborious than other methods.