| A major goal of animal production is to improve reproductive efficiency. However the genetic improvement progress was limited due to its low heritability. Recently, the morden molecular genetics provides a new approach to improve litter size trait. In this study, we analyzed 133 differentially expressed (DE) genes in PMSG-hCG stimulated preovulatory ovarian follicles of Large White and Chinese Erhualian sows, and mapped them to the intervals of reproduction QTLs. qRT-PCR were used to validate eight differentially expressed genes. According to the position and biological function of these DE genes, we choose CTNNAL1, Wnt-10b, BMPR-IB, WIF-1 as candidate genes and then detected SNPs, the association analysis had been done between the polymorphisms and the reproductive performance in Large White and DIV populations. The main results are as follows:1. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses of 95 annotated DE gene lists were carried out. GO annotation mapping revealed that the genes assigned GO terms for regulation of cellular process, biological regulation, developmental process, cell communication and signal transduction and so on. Through BLAST of the 133 probes sets against Ensembl Sus_scrofa database (Sscrofa9, Apr 2009),116 differentially expressed probes sets (transcripts) were in silico mapped pig chromosome covering 1-18 and X. Ninety-two DE transcripts were located to porcine reproduction QTL chromosomal regions.2. The expression pattern of 8 genes including GALP, ITM2A, CTNNBIP1, WIF1, BMPR-IB, CTNNAL1, LOC396850, CNN2 were validated by fluorescence quantitative real time polymerase chain reaction (RT-PCR). The Pearson correlation coefficients of the relative expreesion level of above genes between microarry and qRT-PCR were 0.790, 0.629,0.929,0.950,0.615,0.820,0.614, and-0.766 respectively.3. CTNNAL1 Gene:(1) A 2197bp cDNA sequence of pig CTNNAL1 gene was isolated, including the complete sequence of exons 2-17, partial sequence of exon 1 and exon 18; A 416bp DNA sequence of CTNNAL1 gene was isolated. (2) a PCR-AluI-RFLP assay was established to detect the C/G mutation in the exon 15 and allele G was the dominant allele in all tested pig populations. (3) Association analysis for the C/G with reproductive traits was preformed in two independent populations. The statistical analysis results showed that:In Large White pig population, AluI polymorphism was associated with NBA (P＜0.01), the additive effect value was 0.54±0.20 (P＜0.01); In DIV pig population, AluI polymorphism was associated with TNB and NBA (P<0.05), the additive effect value was 1.40±0.46 and 1.23±0.46 (P<0.01).4. Wnt-10b Gene:(1) The statistical analysis results showed that:allele C was the dominant gene. (2) Association analysis for the C/T with reproductive traits was preformed in two independent populations. In Large White pig first parity group, Sacll polymorphism was associated with NBA (P<0.05).5. WIF-1 Gene:(1) A 1282bp cDNA sequence of pig WIF-1 gene was isolated, including exons 1-9. (2) A 667bp DNA sequence of pig WIF-1 gene in the intron 2 was isolated, and many potential mutations were detected by comparative sequencing.6. BMPR-IB Gene:(1) The statistical analysis results showed that:allele C was the dominant gene. (2) Association analysis for the C/G with reproductive traits was preformed in two independent populations. In DIV pig population, Sacll polymorphism was associated with TNB (P<0.1). |
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