Optimization of transfection efficiency of primary dermis derived fibroblasts of sheep

Recently CRISPR Cas based gene editing technology has come to limelight. The technology has been used to produce gene edited animals by editing desired genes in primary fibroblasts and use them as nuclear donors in SCNT procedures. The editing machinery is usually delivered into cultured cells in vector format. Owing to hardy nature of primary cells to transfection, here we have compared two common transfection strategies viz. chemical and electroporation to assess transfection efficiencies. The vector used was4.7 Kb sized EGFPC1(enhanced green fluoroscent protein), harbouring green fluoroscent protein (GFP) marker. In chemical based strategy we used cationic polymer based chemical reagent for transfection and achieved the highest transfection efficiency of 20% while as in electroporation strategy we were able to achieve 70% transfection efficiency with good cell viability. Furthermore, electrotransfected cells had good morphological and proliferative characteristics as compared to chemically transfected cells.