Improving rates of germline transmission in chimeric chickens using busulfan solubilised sustained-release emulsion.

The administration of a busulfan solubilised sustained-release emulsion was used to improve the efficiency of endogenous primordial germ cell (PGC) depletion in early chicken embryos. Busulfan was solubilised in N,N-dimethylformamide and diluted 10-fold in phosphate-buffered saline [PBS(-)]. The sustained-release emulsion was prepared by mixing equal amounts of busulfan solubilised solution and sesame oil using a filter. The appropriate time and dose of busulfan sustained-release emulsion administration were established to optimise our novel germline replacement technique. Injecting busulfan sustained-release emulsion into the yolk of recipient embryos at zero hours of incubation did not disturb the movement and proliferation of donor cells during the generation of germline chimeras. A dose of 100 μg of busulfan resulted in the greatest reduction of endogenous PGCs in embryonic gonads and the most successful hatchability of sterilised embryos. Test-cross analysis revealed that the germline transmission rate in busulfan treated chickens was significantly higher than in non-treated controls (99.5 versus 6.0%). This unique and efficient germline transmission system has potential applications in the conservation of endangered and y or rare avian species. [ABSTRACT FROM AUTHOR]