The transcription factor Doublesex and Mab-3 Related Transcription Factor 1 (DMRT1) is essential for testis differentiation and male fertility, but less is known about its potential involvement in ovarian differentiation. In mice, Dmrt1-/- females are fertile, although they show a reduction of Stra8 expression. Using our DMRT1 mutant rabbit model, we analyzed ovarian differentiation when DMRT1 expression is abolished. First, we observed that during early gonadal development, DMRT1-/- gonads display a specific transcriptome different from control ovaries and control testes. Second, we assessed the infertility phenotype of mutant females. DMRT1 is highly expressed in germ cells until meiosis initiation, and its disruption leads to a down-regulation of key genes involved in the meiotic process. Chromatin immunoprecipitation sequencing data highlighted that DMRT1 can bind these genes in the rabbit ovary. Our study using the rabbit model shows that in addition to its role in testicular differentiation, DMRT1 has two important functions in ovarian differentiation: an initial involvement in the differentiation of somatic cells and a pivotal role in the differentiation of germ cells. Our data suggested that DMRT1 could be considered in human health as a gene necessary for female fertility.