Time-course and reference-based transcriptome analysis of the immature gonad of the sterlet Acipenser ruthenus

In this study, we performed RNA sequencing of the immature gonads of the sterlet Acipenser ruthenus to identify their sex-biased genes at the onset of sexual differentiation. Overall design: Fish from a single batch were sampled at 4-week intervals from 4 to 28 weeks post hatching (wph). Gonads were individually homogenized with a pestle in ice-cold TRI Reagent. Due to their small size, gonads including surrounding tissues such as Wolffian ducts were excised at 4 wph, while gonads without these tissues were collected at 8-28 wph. After pooling the three gonadal homogenates from the same wph and genetic sex, total RNA was extracted from each of the 28 pools. After the quality and quantity assessment, mRNA was purified from the total RNA. The multiplexed sequencing library was prepared using MGIEasy RNA Directional Library Prep Set (MGI Tech, Shenzhen, China). After equimolar pooling of the indexed libraries, sequencing was performed using DNBSEQ-G400RS with High-Throughput Sequencing Set (MGI Tech) for paired-end, 150 bp reads.