Project description:An F2 cross between two highly divergent porcine breeds for most productive traits, including prolificacy, was generated. F2 sows were classified as of high or low prolificacy based on phenotypic records of four consecutive parities (total number of piglets born and number of piglets born alive were recorded). At the fifth gestation, sows were slaughtered at day 30-32 of gestation, and the number of corpora lutea and number of embryos were registered. Samples from different tissues were snap frozen in liquid nitrogen for further studies. Uterus samples were used for total RNA extraction and hybridized on the affymetrix porcine genechip for comparison between high and low prolificacy F2 sows.
Project description:Previous results from a genome scan in a F2 Iberian by Meishan intercross showed several chromosome regions associated with litter size traits. In order to identify candidate genes underlying these QTL we have performed an ovary gene expression analysis during pregnancy. F2 sows were ranked by their estimated breeding values for prolificacy, the six sows with higher EBV (HIGH prolificacy) and the six with lower EBV (LOW prolificacy) were selected. Samples were hybridized to Affymetrix porcine expression microarrays. The statistical analysis with a mixed-model approach identified 221 differentially expressed probes, representing 189 genes. These genes were functionally annotated in order to identify the genetic pathways overrepresented. Among the most represented functional groups the first one was immune system response activation against external stimulus. The second group was made up of genes which regulate the maternal homeostasis by complement and coagulation cascades. The last group was involved on lipid and fatty acid enzymes of metabolic processes, which participate in steroidogenesis pathway. In order to identify powerful candidate genes for prolificacy, the second approach of this study was merging microarray data with position information of QTL affecting litter size, previously detected in the same experimental cross. According to this, we have identified 27 differentially expressed genes co-localized with QTL for litter size traits, which fulfill the biological, positional and functional criteria. Twelve ovary samples: six high prolific sows and low prolific sows, slaughtered after 30 days of pregnancy. Each sample is the average between right and left ovary from each sow.
Project description:Prolificacy related traits are of great economical interest in the pig industry. microRNAs (miRNAs) are post-transcriptional regulators of gene expression important for reproductive processes. In pigs, the roles of ovarian miRNAs during gestation remain unknown although the ovaries are essential during gestation. It has been hypothesised that ovarian miRNAs could participate during the porcine gestation and, moreover, they could influence the prolificacy levels of sows. The miRNA expression profile was compared in the ovaries of pregnant Iberian x Meishan F2 sows displaying extreme phenotypes regarding prolificacy levels defined as the number of embryos (NE) attached to the uterus at 30-32 days of gestation. miR-146a-5p and miR-142-3p were differentially expressed between high (NEM-bM-^IM-%13) and low (NEM-bM-^IM-$11) prolificacy sows. In silico functional analyses of the predicted mRNA targets for these miRNAs revealed that miR-146a-5p targets were mainly involved in the immune system response important for the establishment of the maternal-foetal tolerance, implantation and maintenance of pregnancy. On the other hand, miR-142-3p targets participated in different biological processes that would contribute to the homeostasis maintenance to ensure a correct functional development of the ovaries. miRNAs associated with prolificacy levels could regulate negatively, by a novel post-transcriptional mechanism, their predicted mRNA targets, PPM1K, TLR1 and CPEB2 which have been reported as differentially expressed in the ovaries of pregnant sows regarding the prolificacy levels. Furthermore, among predicted mRNA targets for miRNAs associated with prolificacy, four genes, differentially expressed in the ovaries of pregnant sows regarding prolificacy levels, (LRRK1, BAT1, CPEB2, CCL8) are proposed to be good candidate genes for litter size due to their location within confidence intervals for prolificacy QTL described previously. Overall, it is suggested that the up-regulation of miR-146a-5p and miR-142-3p in the ovaries of pregnant sows could help in the establishment of a uterine environment, which would favor the embryonic development. Total RNA was isolated from uterus of Iberian x Meishan F2 pregnant sows divided into two groups: High prolificacy sows (n=3) and Low prolificacy sows (n=3). RNA was labeled with the Cy3-like Hy3M-bM-^DM-" dye, mixed with a pool of RNA from the six samples labeled with the Cy5-like Hy5M-bM-^DM-" dye, and hybridized to two-color miRCURYM-bM-^DM-" arrays from ExiqonM-BM-..
Project description:Prolificacy related traits are of great economical interest in the pig industry. microRNAs (miRNAs) are post-transcriptional regulators of gene expression important for reproductive processes. In pigs, the roles of ovarian miRNAs during gestation remain unknown although the ovaries are essential during gestation. It has been hypothesised that ovarian miRNAs could participate during the porcine gestation and, moreover, they could influence the prolificacy levels of sows. The miRNA expression profile was compared in the ovaries of pregnant Iberian x Meishan F2 sows displaying extreme phenotypes regarding prolificacy levels defined as the number of embryos (NE) attached to the uterus at 30-32 days of gestation. miR-146a-5p and miR-142-3p were differentially expressed between high (NE≥13) and low (NE≤11) prolificacy sows. In silico functional analyses of the predicted mRNA targets for these miRNAs revealed that miR-146a-5p targets were mainly involved in the immune system response important for the establishment of the maternal-foetal tolerance, implantation and maintenance of pregnancy. On the other hand, miR-142-3p targets participated in different biological processes that would contribute to the homeostasis maintenance to ensure a correct functional development of the ovaries. miRNAs associated with prolificacy levels could regulate negatively, by a novel post-transcriptional mechanism, their predicted mRNA targets, PPM1K, TLR1 and CPEB2 which have been reported as differentially expressed in the ovaries of pregnant sows regarding the prolificacy levels. Furthermore, among predicted mRNA targets for miRNAs associated with prolificacy, four genes, differentially expressed in the ovaries of pregnant sows regarding prolificacy levels, (LRRK1, BAT1, CPEB2, CCL8) are proposed to be good candidate genes for litter size due to their location within confidence intervals for prolificacy QTL described previously. Overall, it is suggested that the up-regulation of miR-146a-5p and miR-142-3p in the ovaries of pregnant sows could help in the establishment of a uterine environment, which would favor the embryonic development.
Project description:To optimize the genome annotation, nine tissue and one pool RNA libraries (i.e. heart, liver, spleen, lung, kidney, muscle, fat, ovary, pool.) were constructed using the Illumina mRNA-spleeneq Prep Kit
Project description:Previous results from a genome scan in a F2 Iberian by Meishan intercross showed several chromosome regions associated with litter size traits. In order to identify candidate genes underlying these QTL we have performed an ovary gene expression analysis during pregnancy. F2 sows were ranked by their estimated breeding values for prolificacy, the six sows with higher EBV (HIGH prolificacy) and the six with lower EBV (LOW prolificacy) were selected. Samples were hybridized to Affymetrix porcine expression microarrays. The statistical analysis with a mixed-model approach identified 221 differentially expressed probes, representing 189 genes. These genes were functionally annotated in order to identify the genetic pathways overrepresented. Among the most represented functional groups the first one was immune system response activation against external stimulus. The second group was made up of genes which regulate the maternal homeostasis by complement and coagulation cascades. The last group was involved on lipid and fatty acid enzymes of metabolic processes, which participate in steroidogenesis pathway. In order to identify powerful candidate genes for prolificacy, the second approach of this study was merging microarray data with position information of QTL affecting litter size, previously detected in the same experimental cross. According to this, we have identified 27 differentially expressed genes co-localized with QTL for litter size traits, which fulfill the biological, positional and functional criteria.