Long Term in vitro Culture of Chicken Primordial Germ Cells Isolated from Embryonic Blood and Incorporation into Germline of Recipient Embryo

The present study was carried out to develop a long-term in vitro culture system for chicken primordial germ cells (PGCs). PGCs were obtained from the blood of 2.5-day incubated embryos. GFP gene was transferred into the collected PGCs and then cultured on feeder cells derived from the gonads of 7-day incubated embryos. The GFP-positive cells attached loosely to the feeder cells, and their morphology varied from round to fibroblast-like in shape. They proliferated slowly and occasionally formed colonies. The PGCs cultured for 23-61 days were transferred into the bloodstream of recipient embryos, and we examined their incorporation into the germline of chimaeric chickens. Test mating was carried out, and one germline chimaeric chicken was detected out of 28 putative chimaeric chickens. That one chicken was generated by transferring 58-day cultured PGCs and it produced one donor-derived offspring out of 270 examined. Thus, a small percentage of the cultured PGCs retained the ability to migrate to the germinal ridges, thus giving rise to functional gametes, although most of the cultured PGCs differentiated during the culture period. In conclusion, a germline chimaeric chicken was generated by transferring PGCs cultured in vitro for about 2 months; the culture system for PGCs developed in the present study will contribute to the germline manipulation in chickens.