Towards the identification of sex-determining gene(s) in channel catfish

Abstract

Sex is the most fundamental feature in the life of an organism. Studying of sex determination is an important area in animal developmental and evolutionary biology, as well as ecology. In teleost, sex determination mechanism exhibits extraordinary plasticity and diversity with respect to the evolutionary pattern. Catfish have a XY male/XX female sex chromosome system. The exact mechanism of sex determination in catfish is unknown at present. As a first step towards the identification of sex-determining genes in catfish, we performed the first transcriptome-level analysis of the catfish testis using high throughput Illumina sequencing to understand the transcriptome of the catfish testes. Gene ontology and annotation analysis suggested that many of the male-biased genes identified from the analysis were involved in gonadogenesis, spermatogenesis, testicular determination, gametogenesis, gonad differentiation, and possibly sex determination. Our analysis would lay the basis for further follow-up analysis of genes involved in sex determination and differentiation in catfish. To move toward the goal of identification of the Y-specific fragments in channel catfish, we utilized multiple approaches to get the best assembly and scaffolding of X and Y chromosome in catfish and conducted in silico comparative analysis between these two chromosomes. Sequencing of the super-male (YY) sample, the regular male (XY) sample, and BAC clones on the sex chromosome using the Illumina HiSeq 2000 platform followed by assembly by ABySS and scaffolding by SSPACE. Comparative analyses were performed with the rapidly genome aligning software MUMmer. However, by comparing the nucleotide sequences of the X-chromosome and Y-chromosome in detail, no Y-specific sequence was identified. The present work demonstrates that there is no gene deletion/insertion between X and Y chromosome in channel catfish. The position of the putative catfish sex-determining region was defined by assuming that the previously identified SD marker is right in the middle of the two adjacent informative sex-linked markers in both directions with the lowest recombination frequencies. The BAC markers covering this sex-determining region were used as query to BLASTN-searched in the Y-chromosome scaffolds in order to obtain the full coverage of this region. Genes in this SD region were identified using BLASTX-searched in the non-redundant (nr) protein database for annotation. Thus, the SD region is 203 kbp in length, harboring 10 annotated genes. Gene expression in male vs. female embryos at the early gonad differentiation stages will be tested among these candidate genes.