Nanos3 of the frog <italic>Rana rugosa</italic>: Molecular cloning and characterization.

Nanos is expressed in the primordial germ cells (PGCs) and also the germ cells of a variety of organisms as diverse as <italic>Drosophila</italic>, medaka fish, <italic>Xenopus</italic> and mouse. In <italic>Nanos3</italic>‐deficient mice, PGCs fail to incorporate into the gonad and the size of the testis and ovary is thereby dramatically reduced. To elucidate the role of <italic>Nanos</italic> in an amphibian species, we cloned <italic>Nanos3 </italic>cDNA from the testis of the <italic>R</italic>. <italic>rugosa</italic> frog. RT‐PCR analysis showed strong expression of <italic>Nanos3 </italic>mRNA in the testis of adult <italic>R. rugosa</italic> frogs, but expression was not sexually dimorphic during gonadal differentiation. In <italic>Nanos3</italic>‐knockdown tadpoles produced by the CRISPR/Cas9 system, the number of germ cells decreased dramatically in the gonads of both male and female tadpoles before sex determination and thereafter. This was confirmed by three dimensional imaging of wild‐type and <italic>Nanos3</italic> knockdown gonads using serial sections immunostained for Vasa, a marker specific to germ cells. Taken together, these results suggest that Nanos3 protein function is conserved between <italic>R. rugosa</italic> and mouse. [ABSTRACT FROM AUTHOR]