Project description:Purpose: The goal of this study is to compare endothelial small RNA transcriptome to identify the target of OASL under basal or stimulated conditions by utilizing miRNA-seq. Methods: Endothelial miRNA profilies of siCTL or siOASL transfected HUVECs were generated by illumina sequencing method, in duplicate. After sequencing, the raw sequence reads are filtered based on quality. The adapter sequences are also trimmed off the raw sequence reads. rRNA removed reads are sequentially aligned to reference genome (GRCh38) and miRNA prediction is performed by miRDeep2. Results: We identified known miRNA in species (miRDeep2) in the HUVECs transfected with siCTL or siOASL. The expression profile of mature miRNA is used to analyze differentially expressed miRNA(DE miRNA). Conclusions: Our study represents the first analysis of endothelial miRNA profiles affected by OASL knockdown with biologic replicates.
Project description:N6-methyladenosine (m6A) is the most prominent mRNA modification in eukaryotes, and its potential regulatory role has recently been identified in mammals, plants, and yeast. However, how m6A methylation regulates spermatogenesis remains unknown. In this study, cattle-yak testis tissue was used as the experimental material, and the m6A map was generated through preliminary experiments and methylated RNA immunoprecipitation sequencing. Only spermatogonia and Sertoli cells were observed in cattle-yak testis tissue. Experiments examining the expression of methylation-related enzymes and the overall methylation level showed that the methylation level in the testis of the cattle-yak was slightly lower than that of the sexually mature yak, but significantly higher than that of the pre-sexually mature yak. Annotation analysis indicated that differentially methylated peaks were most frequently concentrated in exonic regions, followed by 3'UTR and finally 5'UTR regions. Through enrichment analysis of differentially expressed genes and differentially methylated corresponding genes, GO analysis of T-vs-Y group mainly involved spermatogenesis, including cytoskeleton, actin binding, etc. KEGG analysis showed that the differential genes were mainly enriched in actin cytoskeleton regulation and MAPK signaling pathway. GO analysis of the T-vs-M group mainly involved protein ubiquitination, ubiquitin ligase complexes, ubiquitin-dependent protein catabolism and endocytosis. KEGG analysis mainly involved apoptosis and Fanconi anemia pathways. This study will lay the foundation for elucidating the molecular mechanism of m6A in male sterility of cattle-yak.
Project description:Testis is the most important male reproductive organ, and the integrity of its physiological function is crucial to the successful production of sperm. In this study, the expression profiles of 11 991 and 8 930 cells in testicular tissue of yak and cattle-yak after sexual maturity were established using Single-cell RNA sequencing. The identification results of cell subpopulations and marker genes were analyzed and their possible mechanisms were predicted.
Project description:Purpose: In order to understand the functional significance of sperm transcriptome in stallion fertility, the aim of this study was to generate a detailed body of knowledge about the sperm RNA profile that defines a normal fertile stallion. Methods: The 50 bp single-end ABI SOLiD raw reads were directly aligned with the horse reference sequence EcuCab2 using ABI aligner software (NovoalignCS version 1.00.09, novocraft.com) which uses multiple indexes in the reference genome, identifies candidate alignment locations for each primary read, and allows completion of the alignment. Results: Next generation sequencing (NGS) of total RNA from the sperm of two reproductively normal stallions generated about 70 million raw reads and more than 3 Gb of sequence per sample; over half of these aligned with the EcuCab2 reference genome. Altogether, 19,257 sequence tags with average coverage ?1 (normalized number of transcripts) were mapped in the horse genome. Conclusion: The sequence of stallion sperm transcriptome is an important foundation for the discovery of transcripts of known and novel genes, and non-coding RNAs, thus improving the annotation of the horse genome sequence draft and providing markers for evaluating stallion fertility. Reproductively fertile Stallion sperm transcriptome as revealed by RNA sequencing
Project description:Deep sequencing of mRNA from 6 organs of yak (Bos grunniens) Analysis of ploy(A)+ RNA of brain,heart,liver,lung,spleen, and stomach of yak (Bos grunniens)
Project description:Studying spermatogenesis is the key to understanding the development mechanism of the yak reproductive system. Although N6-methyladenosine (m6A) RNA modification has been reported to regulate spermatogenesis and reproductive function in mammals, the molecular mechanism of m6A in yak testis development and spermatogenesis remains largely unknown. Therefore, we collected testicular tissue from junile and adult yaks, and found that the m6A level significantly increased after sexual maturity in yak. In MeRIP-seq, 1,702 hypermethylated peaks and 724 hypomethylated peaks were identified. The hypermethylated differentially methylated RNAs (DMRs) (CIB2, AK1, FOXJ2, PKDREJ, SLC9A3, and TOPAZ1) mainly regulated spermatogenesis, while the hypomethylated DMRs (BCL6, USP25, CD96, EPHA2, and TAF12) mainly participated in immune response. Functional enrichment analysis showed that DMRs were significantly enriched in the adherens junction, gap junction, and Wnt, PI3K, and mTOR signaling pathways, regulating cell development, spermatogenesis, and testicular endocrine function. The functional analysis of differentially expressed genes showed that they were involved in the biological processes of mitosis, meiosis, and flagellated sperm motility during sexual maturity of yak testis. We also screened the key regulatory factors of testis development and spermatogenesis by combined analysis, which included BRCA1, CREBBP, STAT3, and SMAD4. This study has important research value and practical guiding significance for yak genetic improvement, particularly in understanding the molecular mechanism and evolutionary dynamics of the progression of testis developmental stages in yaks.