Efficiency of Chicken Blastoderm Cell Transfection Combining two Nonviral Methods: Electroporation and Synthetic Carriers

Abstract

KINAL A., KOZ£OWSKA I., £AKOTA P., WAWRZYÑSKA M., KIJEWSKA E. 2017. Efficiency of chicken blastoderm cell transfection combining two nonviral methods: electroporation and synthetic carriers. Folia Biologica (Kraków) 65: 33-41.Blastoderm cells (BCs) are an efficient tool in the generation of chimeric and transgenic chickens. This study includes a comprehensive analysis of the transfection efficiency of BCs using a new method combining electroporation and lipofection. BCs derived from stage X fertilised eggs were subjected to transfection by electroporation and lipofection with the pOVAINT plasmid, which encoded the human interferon alpha 2a. Based on our previous results, the most optimal parameters for electroporation (iso-osmotic solution, two electrical pulses at a voltage of 25 V and a duration of 500 µs) were used. For lipofection, 5 synthetic carriers were tested. Approx. 85.8% of the BCs were successfully transfected when electroporation with a cationic lipid DDAB / DOPE was combined. Injection of transfected cells in vitro into chicken BCs (stage X) of recipient embryos allowed for the introduction of a human interferon alpha-2a gene, the presence of which was detected by PCR, into their genome. Injection efficiency was 65% of the embryos, with a confirmed presence of introduced genes. The presence of the transgene in selected organs (gonads, femoral muscle, brain, pectoral muscle, torso, heart, blood of the embryo depending on the day of incubation) was also demonstrated.