EVALUATION OF TRANSFECTED FIBROBLAST CELLS -OF SAHIWAL CATTLE FOR TRANSFECTION / NUCLEAR TRANSFER STUDIES

Abstract

The development of an efficient transfection system in livestock cells is an important step towards
investigating gene transfer and has impact mainly in agriculture and fundamental biology. The
developmental capacity of transfected fibroblast cells isolated from skin piece of ear pinna of adult
Sahiwal cattle was studied. Effect of transfection and in vitro culture of cells for six successive passages
post transfection was evaluated. Important factors involved in cationic liposome mediated gene transfer
were evaluated through in vitro transfection of bovine fibroblast cells. Transfection of plasmid DNA
complex (pEGFP and DNAfectin reagent) was evaluated and standardized utilizing the parameters
concentration of DNA, lipofection reagent, cell density, and the effect of transfection time on the
efficiency of indigenous Sahiwal cattle fibroblasts to express a reporter gene. Stable transfections
were conducted using a green fluorescence protein (GFP) reporter plasmid containing the neomycin
resistance gene.