PLURIPOTENT CELL CULTURE ENGINEERING AND THE APPLICATION FOR AVIAN BIOTECHNOLOGY: Avian Pluripotent Cell Culture Engineering

摘要

Pluripotent cells have been considered as one of the most useful tools for production of transgenic birds, secretion of Pharmaceuticals in eggs, regeneration of avian organs and conservation of endangered birds via germline chimeras. To produce the germline chimeras, long term culture of the pluripotent cells is essential. Thus, the avian pluripotent cell culture engineering techniques have been one of the most fundamental subjects in avian biotechnology. Recently, a novel strategy has been established in our lab to culture the pluripotent cells derived from center of the area pellucida in stage X blastoderm in chickens. The cultured cells were positively stained with SSEA-1 and Alkaline Phosphatase. Developmental fate of the cultured cells in chimeras was analyzed. The cultured donor cells were microinjected into the subgerminal cavity of the recipients' blastoderm from which cells in the center of the area pellucida were removed. The manipulated embryos were cultured until hatching. Many of the generated chimeras transmitted donor derived gametes to their offspring. It was considered that the cultured cells derived from blastoderm retained the high potency to differentiate into both germ cells and somatic cells. Prior to microinjection, foreigngenes were transfected into the cultured cells. When the chimeric embryos were generated, strong expression of the introduced genes were detected in the embryos. These approaches should open up new frontier for future avian biotechnology.