De novo transcriptome analysis of gonads reveals the sex-associated genes in Chinese hook snout carp Opsariichthys bidens

Abstract

As a small farmed fish, Opsariichthys bidens shows sex phenotypic difference between male and female individuals, but the molecular mechanism of gonadal development is still unclear. This study performed transcriptome sequencing on the adult testis and ovary of O. bidens to identify sex-related genes. A total of 50,321 unigenes were obtained from six libraries of gonadal tissues, with a total length of 69,070,743 bp and an average length of 1372 bp. Moreover, 29,889, 24,874, 14,564, 18,519, 27,195 and 22,558 unigenes were annotated in the non-redundant protein database, Swissprot, Kyoto Encyclopedia of Genes and Genomes (KEGG), eukaryotic orthologous group (KOG), evolutionary genealogy of genes: Non-supervised Orthologous Groups (eggNOG) and Gene Ontology (GO) databases, respectively. By comparing testis and ovary libraries, 12,609 differentially expressed genes were identified, of which 5975 were up-regulated in the testis and 6634 were up-regulated in the ovary. Enrichment analysis of differentially expressed genes resulted in 3119 enriched GO items (padj<0.01) and 47 enriched KEGG pathways (padj<0.01). Subsequently, the expression levels of nine sex-related genes, including dazl, dnd, nanog, piwi, vasa, zp4, amh, cyp19a and dmrt1 were confirmed by qRT-PCR. Furthermore, molecular cloning and in situ hybridization demonstrated that both cyp19a and vasa were conserved in O. bidens. In addition, cyp19a was mainly expressed in early developmental stages of oocytes in the ovary but not in the testis. While vasa was present in both testis and ovary, and was highly expressed in the early stages of gametogenesis in gonads. These findings provide basic support for further research on the function of sex-related genes and the molecular mechanisms of sex determination in O. bidens.