Generation of germ cell-deficient pigs by NANOS3 knockout

NANOS3 is an evolutionarily conserved gene expressed in primordial germ cells that is important for germ cell development. Germ cell deletion by NANOS3 knockout has been reported in several mammalian species, but its function in pigs is unclear. In the present study, we investigated the germline effects of NANOS3 knockout in pigs using CRISPR/Cas9. Embryo transfer of CRISPR/Cas9-modified embryos produced ten offspring, of which one showed wild-type NANOS3 alleles, eight had two mutant NANOS3 alleles, and the other exhibited mosaicism (four mutant alleles). Histological analysis revealed no germ cells in the testes or ovaries of any of the nine mutant pigs. These results demonstrated that NANOS3 is crucial for porcine germ cell production.