GSE136603
Severe sperm acrosome overgrowth and infertility in mice lacking chromosome 18 pachytene piRNA
Purpose: piRNAs are a class of germline-specific, small non-coding RNAs required to maintain genome integrity and preserve RNA homeostasis essential for male gametogenesis. During mouse spermatogenesis, male germ cells transcribe piRNAs at pre-pachytene anad pachytene stages of development from genetically distince loci. The most well-studied and conserved function of pre-pachytene piRNAs is repression of transposons to ensure the integrity of the germline genome. In contrast to pre-pachytene piRNAs, the mode of action and function of pachytene piRNA is much less understood. To investigate and characterize the functional importance of pachytene piRNAs in spermatogenesis and male fertility, we have used CRISPR/Cas9 and established mutant mice which have deleted a bi-directional promoter of the pachytene piRNA cluster on Chr18. Mutant mice have a male sterile  phenotype with acrosome overgrowth and defective motility. Overall design: Methods: Testes mRNA and ncRNA profiles of P28 wild-type (+/+), heterozygous (+/-), and homozygous (-/-) mice were generated by deep sequencing , in triplicate, using Illumina's TruSeq Stranded Total RNA kit with Ribo-Zero following the manufacturer instruction. The fragment size of RNAseq libraries was verified using the Agilent 2100 Bioanalyzer and the concentrations were determined using Qubit instrument. The libraries were loaded onto the Illumina Hiseq 3000 for 2X50 bp paired end read sequencing. The fastq files were generated using the bcl2fastq software for further analysis.