Sex is determined by various transcription factors. Dmrt1, a gene that contains the DM domain (Doublesex/Mab-3 DNA-binding motif), is known to be one of the critical factors required for testicular development in vertebrates. Recently, other DM domain-containing genes (DMRT genes) were isolated in mice and fish. In mice, three of the seven known DMRT genes are expressed in the embryonic gonad. In amphibians, however, only Dmrt1 and Dmrt4 have been cloned. To clarify the role(s) of DMRT genes in sex determination in vertebrates, the molecular cloning of amphibian DMRT genes would be very helpful. In this study, we isolated Dmrt2, -3, and -5 from the frog Rana rugosa and analyzed by RT-PCR their expression during sex determination. The analysis revealed that all three genes are expressed in the developing gonad/mesonephros complex during sex determination with no dimorphism. The results suggest that Dmrt2, -3, and -5 may be involved in sexual development in R. rugosa and that they may function in both testicular and ovarian differentiation.