We identified a sex-linked, recessive body color gene, presently designated w (whitish-yellow), in the frog Rana rugosa from the Iwakuni population in Western Japan. This is the first time a sex-linked body color gene was found in amphibians so far. In this population of R. rugosa, males are the heterogametic sex, but the sex chromosomes are still homomorphic. When heterozygous males (Ww), which were produced by crossing a whitish-yellow female (ww) found in the field and a wild-type male (WW) of the same population, were backcrossed to the homozygous whitish-yellow female (ww), the resultant male offspring were all wild-type, whereas the females were all whitish-yellow. This result definitely indicates that w is recessive and X-linked, and its wild-type allele W is located on the Y chromosome. Using this strain (X(w)X(w) female and X(w)Y(W) male), we found that expression of Dmrt1 and Rspo1, which are involved in testicular and ovarian differentiation in vertebrates, was higher in males and females, respectively, prior to the onset of the sexually dimorphic expression of Cyp17 and Cyp19, which are involved in biosynthesis of sex steroids and are critical markers of gonadal sex differentiation.
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