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Optimization of transfection methods for Huh7 and Vero cells: a comparative study
Availability of an efficient transfection protocol is the first determinant in success of gene transferring studies in mammalian cells which is accomplished experimentally for every single cell type. Herein, we provide data of a comparative study on optimization of transfection condition by electroporation and chemical methods for Huh-7 and Vero cells. Different cell confluencies, DNA/reagent ratios and total transfection volumes were optimized for two chemical reagents including jetPEI™ and Lipofectamine™ 2000. Besides, the effects of electric field strength and pulse length were investigated to improve electroporation efficiency. Transfection of cells by pEGFPN1 vector and tracking the expression of GFP by FACS and Fluorescence Microscopy analysis were the employed methods to evaluate transfection efficiencies. Optimized electroporation protocols yielded 63.73 ± 2.36 and 73.9 ± 1.6 % of transfection in Huh7 and Vero cells respectively, while maximum achieved level of transfection by jetPEI™ was respectively 14.20 ± 0.69 and 28 ± 1.11 % for the same cells. Post transfectional chilling of the cells did not improve electrotransfection efficiency of Huh7 cells. Compared to chemical based reagents, electroporation showed the superior levels of transfection in both cell lines. The presented protocols should satisfy most of the experimental applications requiring high transfection efficiencies of these two cell lines.
Key words: transfection efficiency, Huh-7, Vero, electroporation, optimization
Tsitologiya i Genetika 2012, vol. 46, no. 6, pp. 19-27
E-mail: rfarzin pasteur.ac.ir
|Coded & Designed by Volodymyr Duplij||Modified 02.04.23|