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Background: Toll-like receptors (TLR) are crucial in innate immunity for the recognition of a broad range of microbial pathogens and are expressed in multiple cell types. There are 10 TLR genes described in the pig genome., Results: With a twofold objective i.e. to catalogue genetic variants in porcine TLR genes and develop a genotyping array for genetic association studies on immune-related traits, we combined targeted sub-genome enrichment and high-throughput sequencing to sequence the 10 porcine TLR genes in 266 pigs from 10 breeds and wild boars using a DNA-pooling strategy. We identified 306 single nucleotide variants across the 10 TLR and 11 populations, 87 of which were novel. One hundred and forty-seven positions i.e. six stop-gains and 141 non-synonymous substitutions were predicted to alter the protein sequence. Three positions were unique to a single breed with alternative allele frequencies equal to or higher than 0.5. We designed a genotyping array for future applications in genetic association studies, with a selection of 126 variants based on their predicted impact on protein sequence. Since TLR4, TLR7 and TLR9 were underrepresented in this selection, we also included three variants that were located in the 3'UTR of these genes. We tested the array by genotyping 214 of the 266 sequenced pigs. We found that 93 variants that involved the 10 TLR genes were polymorphic in these animals. Twelve of these variants were novel. Furthermore, seven known variants that are associated with immune-related phenotypes are present on the array and can thus be used to test such associations in additional populations., Conclusions: We identified genetic variations that potentially have an impact on the protein sequence of porcine TLR. A genotyping array with 80 non-synonymous, 10 synonymous and three 3'UTR polymorphisms in the 10 TLR genes is now available for association studies in swine populations with measures on immune-related traits.

Background: In previous studies on an Iberian x Landrace cross, we have provided evidence that supported the porcine ELOVL6 gene as the major causative gene of the QTL on pig chromosome 8 for palmitic and palmitoleic acid contents in muscle and backfat. The single nucleotide polymorphism (SNP) ELOVL6:c.-533C > T located in the promoter region of ELOVL6 was found to be highly associated with ELOVL6 expression and, accordingly, with the percentages of palmitic and palmitoleic acids in longissimus dorsi and adipose tissue. The main goal of the current work was to further study the role of ELOVL6 on these traits by analyzing the regulation of the expression of ELOVL6 and the implication of ELOVL6 polymorphisms on meat quality traits in pigs., Results: High-throughput sequencing of BAC clones that contain the porcine ELOVL6 gene coupled to RNAseq data re-analysis showed that two isoforms of this gene are expressed in liver and adipose tissue and that they differ in number of exons and 3'UTR length. Although several SNPs in the 3'UTR of ELOVL6 were associated with palmitic and palmitoleic acid contents, this association was lower than that previously observed with SNP ELOVL6:c.-533C > T. This SNP is in full linkage disequilibrium with SNP ELOVL6:c.-394G > A that was identified in the binding site for estrogen receptor alpha (ERα). Interestingly, the ELOVL6:c.-394G allele is associated with an increase in methylation levels of the ELOVL6 promoter and with a decrease of ELOVL6 expression. Therefore, ERα is clearly a good candidate to explain the regulation of ELOVL6 expression through dynamic epigenetic changes in the binding site of known regulators of ELOVL6 gene, such as SREBF1 and SP1., Conclusions: Our results strongly suggest the ELOVL6:c.-394G > A polymorphism as the causal mutation for the QTL on pig chromosome 8 that affects fatty acid composition in pigs.

Background: Fat content and fatty acid composition in swine are becoming increasingly studied because of their effect on sensory and nutritional quality of meat. A QTL (quantitative trait locus) for fatty acid composition in backfat was previously detected on porcine chromosome 8 (SSC8) in an Iberian x Landrace F2 intercross. More recently, a genome-wide association study detected the same genomic region for muscle fatty acid composition in an Iberian x Landrace backcross population. ELOVL6, a strong positional candidate gene for this QTL, contains a polymorphism in its promoter region (ELOVL6:c.-533C < T), which is associated with percentage of palmitic and palmitoleic acids in muscle and adipose tissues. Here, a combination of single-marker association and the haplotype-based approach was used to analyze backfat fatty acid composition in 470 animals of an Iberian x Landrace F2 intercross genotyped with 144 SNPs (single nucleotide polymorphisms) distributed along SSC8., Results: Two trait-associated SNP regions were identified at 93 Mb and 119 Mb on SSC8. The strongest statistical signals of both regions were observed for palmitoleic acid (C16:1(n-7)) content and C18:0/C16:0 and C18:1(n-7)/C16:1(n-7) elongation ratios. MAML3 and SETD7 are positional candidate genes in the 93 Mb region and two novel microsatellites in MAML3 and nine SNPs in SETD7 were identified. No significant association for the MAML3 microsatellite genotypes was detected. The SETD7:c.700G > T SNP, although statistically significant, was not the strongest signal in this region. In addition, the expression of MAML3 and SETD7 in liver and adipose tissue varied among animals, but no association was detected with the polymorphisms in these genes. In the 119 Mb region, the ELOVL6:c.-533C > T polymorphism showed a strong association with percentage of palmitic and palmitoleic fatty acids and elongation ratios in backfat., Conclusions: Our results suggest that the polymorphisms studied in MAML3 and SETD7 are not the causal mutations for the QTL in the 93 Mb region. However, the results for ELOVL6 support the hypothesis that the ELOVL6:c.-533C > T polymorphism has a pleiotropic effect on backfat and intramuscular fatty acid composition and that it has a role in the determination of the QTL in the 119 Mb region.