Optimization of transfection system of goat fetal fibroblast cells mediated by Lipofectamin~(TM) LTX

To prepare donor cells for somatic cell nuclear transplantation and produce transgenic goat,a method of foreign gene efficiently transfecting into goat fetal fibroblast cells(gFFCs) was established.Using GFP as a reported gene,four factors including the number of inoculated cells,dose of plasmid DNA,ratio of liposome to plasmid DNA and exposure time of the cells to DNA-liposome complexes were investigated and optimized by LipofectaminTM LTX and PLUSTM reagent.After 24 h,transfection efficiency was evaluated as the percentage of transfected cells under fluorescence microscope.The results showed that the gFFCs cultured in 24 wells with 6×104 cells per well,transfected 24 h later,0.6 μg plasmid DNA per well,and ratio of liposome to plasmid DNA 4.5∶1.0,and exposure of the cells to DNA-liposome complexes for 6 h resulted in the highest transfection efficiency,which was 81.2%.