Sperm transcriptome of Murrah bull
The whole transcriptome Sequencing WTS libraries were prepared using the NEBNext Ultra II Directional RNA library kit. The total 200 ng RNA was prepared for library construction. Prior to library preparation, rRNA depletion was performed using the QIAseq FastSelect kit. The buffalo sperm RNAs were already biologically fragmented, the RNA fragmentation step was modified for partially degraded RNA. Random primers were annealed to the RNA fragments, followed by first-strand synthesis using reverse transcriptase. The complementary strand was then synthesized using DNA polymerase I. The resulting ds-cDNA underwent several enzymatic steps to repair the 3' ends, followed by adapter ligation. The adapter-ligated fragments were purified and enriched via PCR. Post-enrichment, the library was quantified and the size distribution was assessed using the Agilent D5000 ScreenTape System. Finally, the libraries were subjected to 150 bp paired-end RNA sequencing on the Illumina NovaSeq 6000 platform, generating 150 bp paired-end data.