Preliminary Identification Of Pork Water Holding Capacity-Related Genes Using Porcine SNP-Chip

Water holding capacity (WHC) plays a key role in pork quality. To increase WHC level is one of the effective ways to improve pork quality. The aim of this study is to identify novel molecular markers or function genes related to pork WHC using porcine SNP-chip.168 Duroc×Landrace×Yorkshire (D×L×Y) pigs were sampled for WHC determining using out-pressure method.24 out of 168 individules were used for whole-genome associasion analysis with WLR using Illumina Porcine 60K chip. Five candidate SNPs which reside in reported WHC QTLs or possible WHC-related genes were selected for further investigation. For those SNPs being proved to be associated with WLR, the differences of gene expression level among distinct genotypes were assayed. The correlations between candidate gene expression and WLR were analyzed as well.1. Identification of WLR-related SNPs with SNP-chipThe whole-genome association analysis revealed 5520 WHC-related SNPs with F value more than 3,4239 SNPs of which distributed seperately on 18 euchromosomes. The numbers of SNPs harbored in each euchromosome varied from 99 to 544. The Sus Scrofa Chromosome (SSC) 15 had the lowest SNP density of 0.9 per Megabase (Mb), while SSC7 was of the highest density of 2.9 per Mb. The average SNP density across the whole genome was 1.8 per Mb.Five candidate SNPs were selected for the next-step analysis which located in the reported QTLs or the possible candidate genes, of which 3 SNPs reside in KCNN2、FBN2 and ARHGAP26 genes of SSC2 QTL regions,2 SNPs in MUC4 and ITGA-9 genes of SSC13.2. Association analysis of candidate SNPs with WLRFive candidate SNPs were genotyped by PCR-SSCP or AS-PCR in more samples. The association analysis of candidate SNPs with WLR indicated that no significant differences existed among the 3 genotypes of KCNN2 and MUC4 gene (p>0.05). Although there was no difference between the two homozygotes of FBN2 gene, the WLR of AG genotype was higher than that of AA homozygote (p<0.05). There were significant differences of WLR between the two homozygotes for both ARHGAP26 and ITGA-9 genes (p<0.05).3. Correlation of candidate SNPs ARHGAP26 and ITGA-9 mRNA levelThe mRNA expression levels of ARHGAP26 and ITGA-9 genes in longissimus-dorsal muscle tissues were further evaluated by real-time RT-PCR. The results showed that ARHGAP26 mRNA levels of GG genotypes were significantly higher than that of AA genotypes (p<0.05). For ITGA-9, CC genotypes exhibited significantly lower expression level than TT and CT genotypes (p<0.05). Two groups (n=11 for each group) with highest (H) or lowest (L) WLR (p<0.01) were used for the further investigation of the possible correlation of candidate gene expression level with WLR. It is interesting to find both the ARHGAP26 and ITGA-9 gene expression levels were significantly different between the H and L WLR groups (p<0.05). The correlation coefficients (r) were -0.34 and 0.33 for ARHGAP26 and ITGA-9 gene, respectively.