Enhancement of Transfection Efficiency of Fetal Bovine Fibroblast by Manipulation of the Cell Cycle

ORTEGA, NICOLÁS MATÍAS; BARRIONUEVO, BLANCA ELIZABETH; GUIDO, CAROLINA; OLMOS, MARÍA FLORENCIA; FILI, ALEJANDRO; FORCATO, DIEGO; BOSCH, PABLO

Buenos Aires

Jornada; XII Jornadas Anuales de la Sociedad Argentina de Biología; 2010

Sociedad Argentina de Biología

Successful transient and stable transfection of cells represents the first stage in many transgenic strategies based on somatic cell nuclear transfer (SCNT). Therefore, development of more efficient and reliable methods to transfect bovine cells in culture is needed. We studied the effect of the mitotic-blocking agent colchicine on the transient transfection of bovine fetal fibroblasts (BFFs) with a plasmid (pZsGreen1-N1) expressing a green fluorescent reporter protein. Transfection was carried out in presence of different transfection agents: GeneJammer (Stratagene) or FuGene (Roche) in BFF cultures previously treated with increasing colchicine concentrations (0-40 ng/mL) for different incubation periods (3-12 hs). We observed a positive effect of colchicine pretreatment on transfection efficiency at 10 ng/mL for 6 hs as evidenced by a significative increase in the percentage of fluorescent cells. High colchicine concentrations and/or long incubation periods lead to cytotoxicity. Further studies are warranted to define conditions leading to high transfection efficiencies and the karyotypic normalcy of treated cells. Results from this study set the bases to develop improved strategies that will facilitate transfection of BFFs for SCNT transgenesis.