Project description:We sequenced and analyzed the genome of a highly inbred miniature Chinese pig strain, the Banna Minipig Inbred Line (BMI). we conducted whole genome screening using next generation sequencing (NGS) technology and performed SNP calling using Sus Scrofa genome assembly Sscrofa11.1.
Project description:Proteins are essential for sperm function, including fertilizing capacity. Pig spermatozoa, emitted in well-defined ejaculate fractions, clearly vary in their functionality, which would be related to differences in protein composition. This study aimed firstly to update the porcine sperm proteome and secondly to identify proteins differentially expressed among mature spermatozoa from cauda epididymis and those fortuitously delivered in different ejaculate portions. Nine ejaculates from nine mature and fertile boars were manually collected in three separate ejaculate portions: the first 10 mL of sperm-rich ejaculate fraction (SRF), the rest of SRF and the post-SRF. The contents of cauda epididymides of the same boars was collected post-mortem by perfusion. All samples were centrifuged, pooling the resulting sperm pellets within the respective source-sample, which were later split to generate two technical replicates per source. The final eight sperm samples were subjected to iTRAQ-based 2D-LC-MS/MS for protein identification and quantification. A total of 1,723 proteins were identified (974 of Sus Scrofa taxonomy) and 1,602 of them were also quantified (960 of Sus Scrofa taxonomy). After an ANOVA test, 32 Sus Scrofa proteins showed quantitative differences (P < 0.01) among the sperm samples, which were particularly relevant for the functionality of spermatozoa fortuitously ejaculated in the post-SRF. The present study is the first showing quantitative differences in the protein profile of mature spermatozoa, involving proteins clearly implicated in sperm function, that prove the protein profile of boar spermatozoa is remodelled during ejaculation . These findings provide a valuable groundwork for further studies focused on identifying protein biomarkers of sperm fertility.
Project description:Three different stages of pig antral follicles have been studied in a granulosa-cell transcriptome analysis on nylon microarrays (1152 clones). The data have been generated from 7 RNA follicle pools and several technical replicates were made. Four Large, one Medium and two Small follicles pools were considered. For each follicle pool, 2 radioactive labellings were performed. Each membrane was exposed 16 hours (to avoid saturation of the signal of highly expressed genes) and 28 hours (to get some signal from lowly expressed genes). Each probe was hybridised on GPL3971 scag_scai Sus scrofa 1.2K mono array and on GPL3970 scag_scai Sus scrofa 4.6K triplicate array (except for GFS2172 which was labelled only once and hybridised onto 2 GPL3971 scag_scai Sus scrofa 1.2K mono array membranes), so that 4 spots are available for each gene (and 2 spots for GFS2172), for a given RNA and a given radioactive labelling. Keywords: granulosa-cell transcriptome analysis data consisted in (6 RNA x 2 labellings) + (GFS2172 RNA X 1 labelling)= 13 probes, 26 hybridisations, 52 images.
Project description:Long non-coding RNAs (lncRNAs) play important roles in diverse biological processes. However, the landscape of lncRNAs is largely unclear in Sus scrofa. Here we performed stranded RNA-seq on total RNA libraries from over 100 samples of Sus scrofa tissues. We identified 10,813 lncRNAs in Sus scrofa, of which 9,075 are novel. 57% of these lncRNAs were conserved in both human and mouse. These conserved lncRNAs tend to be more tissue-specific than pig-specific lncRNAs, and enriched in reproducible organs (i.e. testis and ovary). We characterized a group of lncRNAs potentially involved in the skeletal muscle development. One such lncRNA, a homolog of maternally expressed gene 3 (MEG3), was specifically expressed in the skeletal muscle at early developmental stage. And its expression pattern is conserved in pig and mouse. By over-expressing and knocking down MEG3 in mouse myoblast cell lines, we demonstrated its novel function as a myoblast proliferation suppressor.
Project description:Three different stages of pig antral follicles have been studied in a granulosa-cell transcriptome analysis on nylon microarrays (1152 clones). The data have been generated from 7 RNA follicle pools and several technical replicates were made. Four Large, one Medium and two Small follicles pools were considered. For each follicle pool, 2 radioactive labellings were performed. Each membrane was exposed 16 hours (to avoid saturation of the signal of highly expressed genes) and 28 hours (to get some signal from lowly expressed genes). Each probe was hybridised on GPL3971 scag_scai Sus scrofa 1.2K mono array and on GPL3970 scag_scai Sus scrofa 4.6K triplicate array (except for GFS2172 which was labelled only once and hybridised onto 2 GPL3971 scag_scai Sus scrofa 1.2K mono array membranes), so that 4 spots are available for each gene (and 2 spots for GFS2172), for a given RNA and a given radioactive labelling. Keywords: granulosa-cell transcriptome analysis
Project description:Copy number variations (CNVs), which represent a significant source of genetic diversity in mammals, are currently being associated with phenotypes of clinical relevance, mostly in humans and mice. Notwithstanding, little is known about the extent of CNV that contributes to genetic variation in farm animals, including pig. This Nimblegen experiment reports a genome-wide high resolution map of copy number variation in the porcine genome. After remapping the initial CNV sequences to the latest genome assembly (Sus scrofa v.9), 84 CNV regions (CNVRs) were identified among the genomes of 21 related porcine samples from Duroc breed. We used a set of NimbleGen CGH arrays that tile across the assayable portion of the pig genome with approximately 2.1 million probes, at a 502 bp average probe spacing (Sus scrofa pre assembly version 6). These CNVRs covered 2 Mb of the genome, and ranged in size from 4 to 352 kb (median size of 12 kb). Together, this analysis provides a useful resource to assist with the assessment of CNVs in the contexts of porcine variation, health and productive efficiency. 21 samples were analyzed in a dye swap loop design. In order to cover the latest, at the time of the experiment, porcine genome assembly (Sus scrofa v.6) with high density, custom Nimblegen HD2 CGH arrays were planned to cover all the chromosomes available with 2.1M probes, which yielded 502 bp of average probe spacing.
Project description:The aim of this study was to gain a principal understanding of alterations within the transcriptome, spliceosome and editome in the pituitary of the domestic pig (Sus scrofa domestica L.), which controls basic physiological processes in the reproductive system, during early pregnancy. In this investigation, we performed extensive analyses of data obtained by high-throughput sequencing of RNA from the gilts' pituitary anterior lobes during embryo implantation and the mid-luteal phase of the estrous cycle, as a stage of the cycle with similar corpus luteum secretory activity to gestation.
Project description:We explored the hypothesis that adipose tissue contains epigenetically distinct subpopulations of adipocytes that are differentially potentiated to record cellular memories of their environment. Adipocytes are large, fragile, and technically difficult to efficiently isolate and fractionate. We developed fluorescence nuclear cytometry (FNC) and fluorescence activated nuclear sorting (FANS) of cellular nuclei from Sus scrofa visceral adipose tissue (SsVAT) using the levels of the pan-adipocyte protein, peroxisome proliferator-activated receptor gamma-2 (PPARg2) to distinguish PPARg2-Positive nuclei from PPARg2-Neg (negative) leukocyte, endothelial, and adipocyte progenitor cell nuclei. PPARg2-Postive VAT nuclei showed 2- to 50-fold higher levels of transcripts encoding most of the chromatin-remodeling factors assayed regulating the methylation of histones and DNA cytosine (e.g., DNMT1, DNMT3A, TET2, TET3, KMT2C, SETDB1, PAXP1, ARID1A, KMT2C, JMJD6, CARM1/PRMT4, PRMT5). PPARg2-Positive nuclei have a large decondensed chromatin structure. TAB-seq demonstrated 5´-hydroxymethylcytosine (5hmC) levels were remarkably dynamic in the gene body of PPARg2-Positive nuclei, dropping 3.8-fold from the highest quintile of expressed genes to the lowest. Sus scrofa VAT (SsVAT) nuclei were isolated from SsVAT. SsVAT nuclei were stained with PPARg2 and sorted with fluorescence activated nuclear sorting (FANS) into PPARg2-High, PPARg2-Med (Medium), PPARg2-Low, and PPARg2-Neg (Negative) four populations.TAB-seq data on 5-hydroxymehtylcytosine (Yu, M. et al. 2012. Cell 149, 1368-1380) was collected from genomic DNA isolated from PPARg2-High, PPARg2-Med+Low (pooled PPARg2-Med and PPARg2-Low), and PPARg2-Neg SsVAT nuclei.