Gene expression in developing goat testes: Sequencing, assembly and identification of caprine spermatogenesis transcriptome

Understanding gene networks necessary to establish and maintain spermatogenesis will provide insight into the development of molecular markers for potential fertility of individual animals. The objective of the current study was to identify candidate genes and gene networks important for the establishment and maintenance of spermatogenesis in the goat testes. We performed a high-throughput RNA sequencing project of caprine testis using Alpine goats as a model system. Sampling included pre-meiotic and meiotic testis parenchyma from 2-mo- and 4-mo-old Alpine bucks (w = 3 animals/time-point). Total RNA isolations were used to generate six single-stranded cDNA libraries, each treated for poly-A selection. In total, the raw reads for tins project represented 436.408.082 sequences reads of an average length of 801 bp. To compare available transcriptomic information of Capris hircus (Yunnan goat) with our high-throughput sequencing project data, we mapped raw reads against publically available goat transcriptome data (ftp://ftp.ncbi.nih.gov/genomes/Capra_hircus). A total of 343.704.188 reads mapped against the reference, and 92.703.894 reads were unmapped. Unmapped reads were used to generate a de novo testes transcriptome assembly of Alpine goat. Mapping information provided insights into the top 1000 highly differentially expressed genes. Results showed an age-specific enrichment for the apoptosis pathway (19 genes; KEGG). Apoptosis is important to establish and maintain spermatogenesis. The expression of 19 apoptosis genes decreased between 2-mo and 4-mo testes. To determine the presence of the apop-tosis gene network in the de novo testes transcriptome. we generated a custom BLAST database and searched 19 genes and 25 variants involved in the apoptosis pathway. BLAST searches and alignment examinations showed evidence for 17 proteins and 20 variants members of the apoptosis pathway. The protein kinase. cAMP-dependent catalytic 3 protein (PRKACB) showed the highest number of variants. Interestingly, tins protein signals cell survival and integrity and is not involved in the execution of cell apoptosis. We have identified gene networks important during pre-meiotic and meiotic phases of spermatogenesis. Future studies will focus on implementing additional bioinformatics approaches to exhaustively and carefully compare completeness of current de novo transcriptome assembly by analysis of additional de novo transcriptome assemblies generated from each individual s