A technical assessment of the porcine ejaculated spermatozoa for a sperm-specific RNA-seq analysis.

A technical assessment of the porcine ejaculated spermatozoa for a sperm-specific RNA-seq analysis. Syst Biol Reprod Med. 2018 Apr 26;:1-13 Authors: Gòdia M, Mayer FQ, Nafissi J, Castelló A, Rodríguez-Gil JE, Sánchez A, Clop A Abstract The study of the boar sperm transcriptome by RNA-seq can provide relevant information on sperm quality and fertility and might contribute to animal breeding strategies. However, the analysis of the spermatozoa RNA is challenging as these cells harbor very low amounts of highly fragmented RNA, and the ejaculates also contain other cell types with larger amounts of non-fragmented RNA. Here, we describe a strategy for a successful boar sperm purification, RNA extraction and RNA-seq library preparation. Using these approaches our objectives were: (i) to evaluate the sperm recovery rate (SRR) after boar spermatozoa purification by density centrifugation using the non-porcine-specific commercial reagent BoviPureTM; (ii) to assess the correlation between SRR and sperm quality characteristics; (iii) to evaluate the relationship between sperm cell RNA load and sperm quality traits and (iv) to compare different library preparation kits for both total RNA-seq (SMARTer Universal Low Input RNA and TruSeq RNA Library Prep kit) and small RNA-seq (NEBNext Small RNA and TailorMix miRNA Sample Prep v2) for high-throughput sequencing. Our results show that pig SRR (~22%) is lower than in other mammalian species and t...
Source: Systems Biology in Reproductive Medicine - Category: Reproduction Medicine Authors: Tags: Syst Biol Reprod Med Source Type: research