Identification and validation of a regulatory mutation upstream of the BMP2 gene associated with carcass length in pigs

Background Carcass length is very important for body size and meat production for swine, thus understanding the genetic mechanisms that underly this trait is of great significance in genetic improvement programs for pigs. Although many quantitative trait loci (QTL) have been detected in pigs, very few have been fine-mapped to the level of the causal mutations. The aim of this study was to identify potential causal single nucleotide polymorphisms (SNPs) for carcass length by integrating a genome-wide association study (GWAS) and functional assays. Results Here, we present a GWAS in a commercial Duroc × (Landrace × Yorkshire) (DLY) population that reveals a prominent association signal (P = 4.49E−07) on pig chromosome 17 for carcass length, which was further validated in two other DLY populations. Within the detected 1 Mb region, the BMP2 gene stood out as the most likely causal candidate because of its functions in bone growth and development. Whole-genome gene expression studies showed that the BMP2 gene was differentially expressed in the cartilage tissues of pigs with extreme carcass length. Then, we genotyped an additional 267 SNPs in 500 selected DLY pigs, followed by further whole-genome SNP imputation, combined with deep genome resequencing data on multiple pig breeds. Reassociation analyses using genotyped and imputed SNP data revealed that the rs320706814 SNP, located approximately 123 kb upstream of the BMP2 gene, was the strongest candidate causal mutation, with a large association with carcass length, with a ~ 4.2 cm difference in length across all three DLY populations (N = 1501; P = 3.66E−29). This SNP segregated in all parental lines of the DLY (Duroc, Large White and Landrace) and was also associated with a significant effect on body length in 299 pure Yorkshire pigs (P = 9.2E−4), which indicates that it has a major value for commercial breeding. Functional assays showed that this SNP is likely located within an enhancer and may affect the binding affinity of transcription factors, thereby regulating BMP2 gene expression. Conclusions Taken together, these results suggest that the rs320706814 SNP on pig chromosome 17 is a putative causal mutation for carcass length in the widely used DLY pigs and has great value in breeding for body size in pigs.

Evidence Against the Causal Relationship Between a Putative Cis-Regulatory Variant of MYH3 and Intramuscular Fat Content in Pigs

Improving meat quality has become the main goal of modern pig breeding. Intramuscular fat content (IMF) is an important trait influencing meat quality of livestock, but the molecular mechanism behind this trait is still unclear. Recently, Cho et al. reported the discovery of the first causal mutation affecting IMF and red flesh color (a*) in pigs, namely XM_013981330.2:g.−1805_−1810del, a 6-bp deletion variant in the porcine MYH3 promoter region. The objective of this study was to reassess the causality of this mutation for its potential commercial application. By Sanger sequencing, we firstly identified several new variants (including a 4-bp deletion) at or near the 6-bp deletion site, which formed four haplotypes in multiple breeds. Unexpectedly, the 6-bp deletion allele, previously determined as the MYH3 Q allele because of its significantly positive effect on IMF and a*, was found not only in Chinese indigenous breeds, but also in four western commercial breeds with relatively lower IMF levels, including Duroc, Large White, Landrace and Pietrain. More surprisingly, we found that the MYH3 Q allele and the haplotypes harboring it had no significant effects on IMF, marbling and color score in three large-scale divergent pig populations: the heterogeneous F6 and F7 pigs and commercial crossbred Duroc × (Landrace × Yorkshire) pigs. Transient transfection analysis in porcine satellite cells showed that the 6-bp deletion variants had a negligible effect on transcription of reporter gene, but could attenuate the MRF (myogenesis regulatory factors)-induced increase in luciferase activity of the MYH3 promoter vector. The MYH3 protein level in muscle did not differ significantly among the haplotype groups. Therefore, our results cannot support the causal relationship between the 6-bp deletion in MYH3 and IMF trait, suggesting that the causal mutation for the IMF QTL on SSC12 needs to be further identified.

BSAseq: an interactive and integrated web-based workflow for identification of causal mutations in bulked F2 populations

Summary With the advance of next-generation sequencing technologies and reductions in the costs of these techniques, bulked segregant analysis (BSA) has become not only a powerful tool for mapping quantitative trait loci but also a useful way to identify causal gene mutations underlying phenotypes of interest. However, due to the presence of background mutations and errors in sequencing, genotyping, and reference assembly, it is often difficult to distinguish true causal mutations from background mutations. In this study, we developed the BSAseq workflow, which includes an automated bioinformatics analysis pipeline with a probabilistic model for estimating the linked region (the region linked to the causal mutation) and an interactive Shiny web application for visualizing the results. We deeply sequenced a sorghum male-sterile parental line (ms8) to capture the majority of background mutations in our bulked F2 data. We applied the workflow to 11 bulked sorghum F2 populations and 1 rice F2 population and identified the true causal mutation in each population. The workflow is intuitive and straightforward, facilitating its adoption by users without bioinformatics analysis skills. We anticipate that the BSAseq workflow will be broadly applicable to the identification of causal mutations for many phenotypes of interest. Availability and implementation BSAseq is freely available on https://www.sciapps.org/page/bsa. Supplementary information Supplementary data are available at Bioinformatics online.

Association of Rs339939442 in the AHR Gene with Litter Size are Inconsistent among Chinese Indigenous Pigs and Western Commercial Pigs

Eastern and Southern Chinese pigs have been imported to Western countries to improve economic traits including fertility in Western pig breeds by intensive selecting Chinese advantage genes. It was reported that the selected Asian-derived non-synonymous mutations including rs339939442 (G > T) in the aryl hydrocarbon receptor (AHR) gene could increase litter size in multiple European commercial lines. The objective of this study is to identify whether rs339939442 in the AHR gene is polymorphic and has an influence on the litter size in 10 pig populations including five Chinese indigenous breeds, one cultivated breed, one lean-type breed, two North American lean-type breeds, and one European lean-type breed. We found that rs339939442 had polymorphism in all 10 populations, whereas rs339939442 was associated with litter size only in French Yorkshire (FRA-Y) and Chinese cultivated Suhuai (SH) pigs containing approximately 75% British Yorkshire pigs ancestry. Our results indicated that rs339939442 in the AHR gene was a potential marker to improve litter size in European commercial lines and the pigs containing ancestries of European commercial lines, whereas this locus maybe not a causal mutation affecting the litter size but only in linkage disequilibrium with the causal mutation for litter size.

Perlman Syndrome

This chapter describes Perlman syndrome, which is an autosomal recessive overgrowth syndrome that presents with a severe phenotype, usually resulting in early postnatal death. Overgrowth is extended to the internal organs, liver, pancreas, and especially kidneys, with histologic findings of focal hamartomas and nephroblastomatosis. These dysplasias predispose to the development of Wilms tumor, a very common occurrence in Perlman syndrome. The condition seems to be very rare, even though mild cases, if any exist, may go undiagnosed. The causal mutation affecting the DIS3L2 gene was discovered only recently and it is not known if this is the only causal gene.