Surrogate production of genome edited sperm from a different subfamily by spermatogonial stem cell transplantation

Abstract

AbstractThe surrogate reproduction technique provides a powerful tool for production of allogenic or xenogeneic gametes derived from endangered species or those with valuable genetic traits. Production of functional donor-derived gametes through intra- or inter-specific spermatogonial stem cell transplantation (SSCT) has been achieved in many species. However, generation of functional gametes from a phylogenetically distant species such as from a different subfamily by SSCT has never been successful. Here, using two small cyprinid fishes, Chinese rare minnow (gobiocypris rarus, for brief:Gr) and zebrafish (danio rerio), which belong to different subfamilies, as donors and recipients for SSCT, we optimized the SSCT technique and successfully obtainedGr-derived sperm carrying targeted genome modifications in zebrafish. We revealed that the transplantedGrspermatogonia supported the host gonadal development and underwent normal spermatogenesis, resulting in a reconstructed fertile testis containingGrspermatids and zebrafish testicular somatic cells. Interestingly, the surrogate spermatozoa resembled those of host zebrafish but not donorGrin morphology and swimming behavior. Finally, we showed thatGr-derived genome edited sperm was successfully produced in zebrafish by cross-subfamily SSCT, when thepou5f3andchdgene knockoutGrSSCs were used as surrogate donors. This is the first report demonstrating the surrogate production of genome edited sperm from a phylogenetically distant species, and this method is feasible to be applied to future breeding of commercial fishes.