Genome-wide association study on chicken carcass traits using sequence data imputed from SNP array

Chicken carcass traits are economically important for the chicken industry. Detecting which genes affect chicken carcass traits is of great benefit to the genetic improvement of this important agricultural species. To investigate the genetic mechanism of carcass traits in chickens, we carried out a genome-wide association study (GWAS). A total of 435 Chinese indigenous chickens were phenotyped for carcass weight (CW), eviscerated weight with giblets (EWG), and eviscerated weight (EW) after slaughter at 91 days and were genotyped using a 600-K single nucleotide polymorphism (SNP) genotyping array. Twenty-four birds were selected for sequencing, and the 600 K SNP panel data were imputed to sequence data with the 24 birds as the reference. Univariate GWASs were performed with GEMMA software using the whole genome sequence data imputed from SNP chip data. Finally, 3, 25, and 63 suggestively significant SNPs were identified to be associated with carcass weight (CW), eviscerated weight with giblets (EWG), and eviscerated weight (EW), respectively. Six candidate genes, RNF219, SCEL, MYCBP2, ETS1, APLP2, and PRDM10 were detected. SCEL and MYCBP2 were potentially associated with these three traits, RNF219 and APLP2 were potentially associated with EWG and EW, and ETS1 and PRDM10 were only potentially associated with EWG and EW, respectively. Compared with forefathers’ research, 10 reported QTLs associated with CW were located within a 5-Mb distance near the SNPs with P value lower than 1×10^?5. This study enriched the knowledge of the genetic mechanisms of chicken carcass traits.     

Impact of variants on type-2 diabetes risk genes identified through genomewide association studies in polycystic ovary syndrome: a case–control study

Polycystic ovary syndrome (PCOS) is a common endocrine disorder in females, and is associated with altered metabolic processes in particular insulin resistance and diabetes mellitus. PCOS shares with type-2 diabetes (T2D) a number of features, including beta cell dysfunction, impaired glucose tolerance and dyslipidaemia. Recently, genomewide association studies (GWAS) have reported a number of genes with reproducible associations and susceptibilities to T2D. To address this, we examined the association between the T2D GWAS candidate genes (CDKAL1, CDKN2B, COL8A1, HHEX, IGF2BP2, KCNJ1, KCNQ1 and SLC30A8) and PCOS in Saudi women. A case–control study, includes 162 cases and 162 controls was enrolled. Genotyping was carried out by the allelic discrimination method. Our results showed that the variants including rs792837 of COL8A1, rs61873498 of KCNQ1 and rs13266634 of SLC30A8 genes to be significantly more frequent in PCOS patients than in controls. Our results suggest that COL8A1, KCNQ1 and SLC30A8, which are previously identified through GWAS as T2D-associated genes, are associated with PCOS.     

Genome-wide association study of <Emphasis Type="Italic">Mycobacterium avium</Emphasis> subspecies <Emphasis Type="Italic">Paratuberculosis</Emphasis> infection in Chinese Holstein

Background

Paratuberculosis is a contagious, chronic and enteric disease in ruminants, which is caused by Mycobacterium avium subspecies paratuberculosis (MAP) infection, resulting in enormous economic losses worldwide. There is currently no effective cure for MAP infection or a vaccine, it is thus important to explore the genetic variants that contribute to host susceptibility to infection by MAP, which may provide a better understanding of the mechanisms of paratuberculosis and benefit animal genetic improvement. Herein we performed a genome-wide association study (GWAS) to identify genomic regions and candidate genes associated with susceptibility to MAP infection in dairy cattle.

Results

Using Illumina Bovine 50?K (54,609 SNPs) and GeneSeek HD (138,893 SNPs) chips, two analytical approaches were performed, GRAMMAR-GC and ROADTRIPS in 937 Chinese Holstein cows, among which individuals genotyped by the 50?K chip were imputed to HD SNPs with Beagle software. Consequently, 15 and 11 significant SNPs (P?<?5?×?10^??5) were identified with GRAMMAR-GC and ROADTDRIPS, respectively. A total of 10 functional genes were in proximity to (i.e., within 1?Mb) these SNPs, including IL4, IL5, IL13, IRF1, MyD88, PACSIN1, DEF6, TDP2, ZAP70 and CSF2. Functional enrichment analysis showed that these genes were involved in immune related pathways, such as interleukin, T cell receptor signaling pathways and inflammatory bowel disease (IBD), implying their potential associations with susceptibility to MAP infection. In addition, by examining the publicly available cattle QTLdb, a previous QTL for MAP was found to be overlapped with one of regions detected currently at 32.5?Mb on BTA23, where the TDP2 gene was anchored.

Conclusions

In conclusion, we identified 26 SNPs located on 15 chromosomes in the Chinese Holstein population using two GWAS strategies with high density SNPs. Integrated analysis of GWAS, biological functions and the reported QTL information helps to detect positional candidate genes and the identification of regions associated with susceptibility to MAP traits in dairy cattle.

     

A genome-wide association study on growth traits in orange-spotted grouper (<Emphasis Type="Italic">Epinephelus coioides</Emphasis>) with RAD-seq genotyping

The orange-spotted grouper, Epinephelus coioides, is one of the most popular fish in China and Southeast Asian countries because of its important economic value. However, molecular mechanism underlying the growth of orange-spotted grouper has never been fully understood. Herein, we performed a genome-wide association study (GWAS) on a natural population of 198 individuals aiming to screen the whole genome of orange-spotted grouper for identification of growth-related loci by restrictionsite associated DNA sequencing. In this research, 261,366 single nucleotide polymorphisms (SNPs) were developed, in which 110 SNPs were identified to be correlated with growth and 20 SNPs were further confirmed to be associated with both body weight and total length. From these identified SNPs, we annotated a total of 34 genes, including adgrb2, csnkza1, cers5, col22a1, creb5, dnd1, dzank1, dnai1, npy2r, fat3, lrrk2, lrp5, map3k9, and so on. Among these candidate genes, npy2r (neuropeptide Y receptor Y2) was reported to play a critical role in growth of the orange-spotted grouper. In addition, population structure, principal component analysis, kinship matrix and linkage disequilibrium were examined to verify the accuracy and reliability of our GWAS results. Our data will also provide a valuable genetic resource for further marker-assisted selection program to improve growth quality in groupers.     

A GWA study reveals genetic loci for body conformation traits in Chinese Laiwu pigs and its implications for human BMI

Pigs share numerous physiological and phenotypic similarities with human and thus have been considered as a good model in nonrodent mammals for the study of genetic basis of human obesity. Researches on candidate genes for obesity traits have successfully identified some common genes between humans and pigs. However, few studies have assessed how many similarities exist between the genetic architecture of obesity in pigs and humans by large-scale comparative genomics. Here, we performed a genome-wide association study (GWAS) using the porcine 60 K SNP Beadchip for BMI and other four conformation traits at three different ages in a Chinese Laiwu pig population, which shows a large variability in fat deposition. In total, 35 SNPs were found to be significant at Bonferroni-corrected 5 % chromosome-wise level (P = 2.13 × 10^?5) and 88 SNPs had suggestive (P < 10^?4) association with the conformation traits. Some SNPs showed age-dependent association. Intriguingly, out of 32 regions associated with BMI in pigs, 18 were homologous with the loci for BMI in humans. Furthermore, five closest genes to GWAS peaks including HIF1AN, SMYD3, COX10, SLMAP, and GBE1 have been already associated with BMI in humans, which makes them very promising candidates for these QTLs. The result of GO analysis provided strong support to the fact that mitochondria and synapse play important roles in obesity susceptibility, which is consistent with previous findings on human obesity, and it also implicated new gene sets related to chromatin modification and Ig-like C2-type 5 domain. Therefore, these results not only provide new insights into the genetic architecture of BMI in pigs but also highlight that humans and pigs share the significant overlap of obesity-related genes.     

SNP markers associated with body size and pelt length in American mink (<Emphasis Type="Italic">Neovison vison</Emphasis>)

Background

Identification of genes underlying production traits is a key aim of the mink research community. Recent availability of genomic tools have opened the possibility for faster genetic progress in mink breeding. Availability of mink genome assembly allows genome-wide association studies in mink.

Results

In this study, we used genotyping-by-sequencing to obtain single nucleotide polymorphism (SNP) genotypes of 2496 mink. After multiple rounds of filtering, we retained 28,336 high quality SNPs and 2352 individuals for a genome-wide association study (GWAS). We performed the first GWAS for body weight, behavior, along with 10 traits related to fur quality in mink.

Conclusions

Combining association results with existing functional information of genes and mammalian phenotype databases, we proposed WWC3, MAP2K4, SLC7A1 and USP22 as candidate genes for body weight and pelt length in mink.

     

Further evidence of the involvement of the Wnt signaling pathway in Dupuytren’s disease

Genetic background plays an important role in the development of Dupuytren’s disease. A genome-wide association study (GWAS) showed that nine loci are associated with the disease, six of which contain genes that are involved in Wnt signaling (WNT2, WNT4, WNT7B, RSPO2, SFRP4, SULF1). To obtain insight in the role of these genes, we performed expression studies on affected and unaffected patient’s tissues. Surgically obtained nodules and cords from eight Dupuytren’s patients were compared to patient-matched control tissue (unaffected transverse palmar fascia). The Wnt-related genes found in the GWAS, the classical Wnt-downstream protein β-catenin, as well as (myo)fibroblast markers were analyzed using real-time qPCR and immunohistochemical stainings for mRNA levels and protein levels, respectively. The collagen-coding genes COL1A1 and COL3A1 were highly upregulated on mRNA level, both in cords and nodules. Three Wnt-related genes were found to be differently regulated compared to control tissue: WNT2 was downregulated in nodules, WNT7B was upregulated in nodules, and SFRP4 was upregulated in nodules and cords. Immunohistochemistry revealed significantly less staining of Wnt2 in cords, but significantly more staining for Wnt7b in nodules. There was significantly more staining of α-SMA in nodules and cord and β-catenin in nodules than in control tissue. We found differences in expression, both at mRNA and protein level, in several Wnt-related genes found earlier to be associated with Dupuytren’s disease. Of these, Wnt7b was upregulated and found in close association with both α-SMA and β-catenin expressing cells, making it a candidate pro-fibrotic mediator in Dupuytren’s disease.     

Search for genetic markers of climatic adaptation in populations of North Eurasia

Genetic diversity of native populations of North Eurasia is investigated using a panel of genetic markers of candidate genes for cold climate adaptation. A high level of within- and between-population variability is detected. Comparative analysis of data on North Eurasian populations combined with data on worldwide populations from the 1000 Genomes and HDGP projects reveals correlations of genetic diversity in candidate genes for cold climate adaptation with key climate parameters, as well as the increase of genetic diversity in markers of this group of genes with the increase of latitude, that is, as modern humans migrated out of Africa. Using the method of searching for extreme empirical values of the coefficient of genetic diversity, signals of directional selection for markers of six genes adaptive to cold (MYOF, LONP2, IFNL4, MKL1, SLC2A12, and CPT1A) are found. The data are discussed in framework of the hypothesis of decanalization of genome–phenome relationships under the pressure of natural selection during human settlement throughout the world.     

A study of candidate genes for day blindness in the standard wire haired dachshund

Background

A genetic study was performed to identify candidate genes associated with day blindness in the standard wire haired dachshund. Based on a literature review of diseases in dogs and human with phenotypes similar to day blindness, ten genes were selected and evaluated as potential candidate genes associated with day blindness in the breed.

Results

Three of the genes, CNGB3, CNGA3 and GNAT2, involved in cone degeneration and seven genes and loci, ABCA4, RDH5, CORD8, CORD9, RPGRIP1, GUCY2D and CRX, reported to be involved in cone-rod dystrophies were studied. Polymorphic markers at each of the candidate loci were studied in a family with 36 informative offspring. The study revealed a high frequency of recombinations between the candidate marker alleles and the disease.

Conclusion

Since all of the markers were at the exact position of the candidate loci, and several recombinations were detected for each of the loci, all ten genes were excluded as causal for this canine, early onset cone-rod dystrophy. The described markers may, however, be useful to screen other canine resource families segregating eye diseases for association to the ten genes.

     

Discovery and characterization of two new stem rust resistance genes in <Emphasis Type="Italic">Aegilops sharonensis</Emphasis>

Key message

We identified two novel wheat stem rust resistance genes, Sr-1644-1Sh and Sr-1644-5Sh in Aegilops sharonensis that are effective against widely virulent African races of the wheat stem rust pathogen.

Abstract

Stem rust is one of the most important diseases of wheat in the world. When single stem rust resistance (Sr) genes are deployed in wheat, they are often rapidly overcome by the pathogen. To this end, we initiated a search for novel sources of resistance in diverse wheat relatives and identified the wild goatgrass species Aegilops sharonesis (Sharon goatgrass) as a rich reservoir of resistance to wheat stem rust. The objectives of this study were to discover and map novel Sr genes in Ae. sharonensis and to explore the possibility of identifying new Sr genes by genome-wide association study (GWAS). We developed two biparental populations between resistant and susceptible accessions of Ae. sharonensis and performed QTL and linkage analysis. In an F₆ recombinant inbred line and an F₂ population, two genes were identified that mapped to the short arm of chromosome 1S^sh, designated as Sr-1644-1Sh, and the long arm of chromosome 5S^sh, designated as Sr-1644-5Sh. The gene Sr-1644-1Sh confers a high level of resistance to race TTKSK (a member of the Ug99 race group), while the gene Sr-1644-5Sh conditions strong resistance to TRTTF, another widely virulent race found in Yemen. Additionally, GWAS was conducted on 125 diverse Ae. sharonensis accessions for stem rust resistance. The gene Sr-1644-1Sh was detected by GWAS, while Sr-1644-5Sh was not detected, indicating that the effectiveness of GWAS might be affected by marker density, population structure, low allele frequency and other factors.

     

Genome-wide association study of heading and flowering dates and construction of its prediction equation in Chinese common wheat

Heading date is one of the most important traits in wheat breeding as it affects adaptation and yield potential. A genome-wide association study (GWAS) using the 90 K iSelect SNP genotyping assay indicated that a total of 306 loci were significantly associated with heading and flowering dates in 13 environments in Chinese common wheat from the Yellow and Huai wheat region. Of these, 105 loci were significantly correlated with both heading and flowering dates and were found in clusters on chromosomes 2, 5, 6, and 7. Based on differences in distribution of the vernalization and photoperiod genes among chromosomes, arms, or block regions, 13 novel, environmentally stable genetic loci were associated with heading and flowering dates, including RAC875_c41145_189 on 1DS, RAC875_c50422_299 on 2BL, and RAC875_c48703_148 on 2DS, that accounted for more than 20% phenotypic variance explained (PVE) of the heading/flowering date in at least four environments. GWAS and t test of a combination of SNPs and vernalization and photoperiod alleles indicated that the Vrn-B1, Vrn-D1, and Ppd-D1 genes significantly affect heading and flowering dates in Chinese common wheat. Based on the association of heading and flowering dates with the vernalization and photoperiod alleles at seven loci and three significant SNPs, optimal linear regression equations were established, which show that of the seven loci, the Ppd-D1 gene plays the most important role in modulating heading and flowering dates in Chinese wheat, followed by Vrn-B1 and Vrn-D1. Additionally, three novel genetic loci (RAC875_c41145_189, Excalibur_c60164_137, and RAC875_c50422_299) also show important effect on heading and flowering dates. Therefore, Ppd-D1, Vrn-B1, Vrn-D1, and the novel genetic loci should be further investigated in terms of improving heading and flowering dates in Chinese wheat. Further quantitative analysis of an F₁₀ recombinant inbred lines population identified a major QTL that controls heading and flowering dates within the Ppd-D1 locus with PVEs of 28.4% and 34.0%, respectively; this QTL was also significantly associated with spike length, peduncle length, fertile spikelets number, cold resistance, and tiller number.     

Genome-wide association study of body weight in Wenshang Barred chicken based on the SLAF-seq technology

Chicken body weight (BW) is an economically important trait, and many studies have been conducted on genetic selection for BW. However, previous studies have detected functional chromosome mutations or regions using gene chips. The present study used the specific-locus amplified fragment sequencing (SLAF-seq) technology to perform a genome-wide association study (GWAS) on purebred Wengshang Barred chicken. A total of 1,286,715 single-nucleotide polymorphisms (SNPs) were detected, and 175,211 SNPs were selected as candidate SNPs for genome-wide association analysis using TASSEL general linear models. Six SNP markers reached genome-wide significance. Of these, rs732048524, rs735522839, rs738991545, and rs15837818 were significantly associated with body weight at 28 days (BW28), while rs314086457 and rs315694878 were significantly associated with BW120. These SNPs are close to seven genes (PRSS23, ME3, FAM181B, NABP1, SDPR, TSSK6L2, and RBBP8). Moreover, 24 BW-associated SNPs reached “suggestive” genome-wide significance. Of these, 6, 13, 1, and 4 SNPs were associated with BW28, BW56, BW80, and BW120, respectively. These results would enrich the studies on BW and promote the use of Chinese chicken, especially the Wenshang Barred chicken.     

Identification and validation of quantitative trait loci controlling seed isoflavone content across multiple environments and backgrounds in soybean

Soybean is important throughout the world not only due to the high seed protein and oil but also owing to the seed isoflavone. To improve the isoflavone concentration in seeds, detecting and mining the stable and reliable quantitative trait loci (QTLs) and related genes in multiple environments and genetic backgrounds become more and more important. In view of this, a F_6:7 recombinant inbred line (RIL) population of 345 lines derived from a cross between Zheng 92116 and Liaodou14 (ZL) was genotyped using 1739 polymorphic SNP and 127 SSR markers in this study and was phenotyped for individual and total seed isoflavone in four environments over 2 years. In total, 48 additive QTLs, which explained 3.00–29.83% of seed isoflavone variation, were identified. Of them, eight QTLs (qDA1_1, qGA1_1, qTIA1_1, qDA1_2, qGA1_2, qTIA1_2, qDA1_3, qTIA1_3) with phenotypic variation explained (PVE) ranging from 14.09 to 28.59% for daidzin, genistin, and total isoflavone were located on the same region of linkage group (LG) A1. These QTLs were further verified in another RIL population derived from Zheng 92116 × Qihuang 30 (ZQ). Meanwhile, the other four overlapping QTLs on linkage group B1, which were associated with glycitin content (qGLB1_1, qGLB1_2, qGLB1_3, qGLB1_4) and explained 16.52 to 29.83% of phenotypic variation, were also verified using the ZQ population. Moreover, the individuals with different genotypes at the common flanking SNP markers for these QTLs on LGs A1 and B1 in the two mapping populations showed significant different isoflavone content, which further validate the QTL mapping results. And also, some candidate genes might participate in the isoflavone biosynthesis processes were found in these stable QTL regions. Thus, the novel and stable QTLs identified and verified in this study could be applied in marker-assisted selection breeding or map-based candidate genes cloning in soybean seed isoflavone genetic improvement in future.     

Next-generation sequencing to identify candidate genes and develop diagnostic markers for a novel Phytophthora resistance gene, <Emphasis Type="Italic">RpsHC18</Emphasis>, in soybean

Key message

A novel Phytophthora sojae resistance gene RpsHC18 was identified and finely mapped on soybean chromosome 3. Two NBS–LRR candidate genes were identified and two diagnostic markers of RpsHC18 were developed.

Abstract

Phytophthora root rot caused by Phytophthora sojae is a destructive disease of soybean. The most effective disease-control strategy is to deploy resistant cultivars carrying Phytophthora-resistant Rps genes. The soybean cultivar Huachun 18 has a broad and distinct resistance spectrum to 12 P. sojae isolates. Quantitative trait loci sequencing (QTL-seq), based on the whole-genome resequencing (WGRS) of two extreme resistant and susceptible phenotype bulks from an F_2:3 population, was performed, and one 767-kb genomic region with ΔSNP-index ≥ 0.9 on chromosome 3 was identified as the RpsHC18 candidate region in Huachun 18. The candidate region was reduced to a 146-kb region by fine mapping. Nonsynonymous SNP and haplotype analyses were carried out in the 146-kb region among ten soybean genotypes using WGRS. Four specific nonsynonymous SNPs were identified in two nucleotide-binding sites–leucine-rich repeat (NBS–LRR) genes, RpsHC18-NBL1 and RpsHC18-NBL2, which were considered to be the candidate genes. Finally, one specific SNP marker in each candidate gene was successfully developed using a tetra-primer ARMS-PCR assay, and the two markers were verified to be specific for RpsHC18 and to effectively distinguish other known Rps genes. In this study, we applied an integrated genomic-based strategy combining WGRS with traditional genetic mapping to identify RpsHC18 candidate genes and develop diagnostic markers. These results suggest that next-generation sequencing is a precise, rapid and cost-effective way to identify candidate genes and develop diagnostic markers, and it can accelerate Rps gene cloning and marker-assisted selection for breeding of P. sojae-resistant soybean cultivars.

     

Gene expression profiles in liver of pigs with extreme high and low levels of androstenone

Background

Boar taint is the unpleasant odour and flavour of the meat of uncastrated male pigs that is primarily caused by high levels of androstenone and skatole in adipose tissue. Androstenone is a steroid and its levels are mainly genetically determined. Studies on androstenone metabolism have, however, focused on a limited number of genes. Identification of additional genes influencing levels of androstenone may facilitate implementation of marker assisted breeding practices. In this study, microarrays were used to identify differentially expressed genes and pathways related to androstenone metabolism in the liver from boars with extreme levels of androstenone in adipose tissue.

Results

Liver tissue samples from 58 boars of the two breeds Duroc and Norwegian Landrace, 29 with extreme high and 29 with extreme low levels of androstenone, were selected from more than 2500 individuals. The samples were hybridised to porcine cDNA microarrays and the 1% most significant differentially expressed genes were considered significant. Among the differentially expressed genes were metabolic phase I related genes belonging to the cytochrome P450 family and the flavin-containing monooxygenase FMO1. Additionally, phase II conjugation genes including UDP-glucuronosyltransferases UGT1A5, UGT2A1 and UGT2B15, sulfotransferase STE, N-acetyltransferase NAT12 and glutathione S-transferase were identified. Phase I and phase II metabolic reactions increase the water solubility of steroids and play a key role in their elimination. Differential expression was also found for genes encoding 17beta-hydroxysteroid dehydrogenases (HSD17B2, HSD17B4, HSD17B11 and HSD17B13) and plasma proteins alpha-1-acid glycoprotein (AGP) and orosomucoid (ORM1). 17beta-hydroxysteroid dehydrogenases and plasma proteins regulate the availability of steroids by controlling the amount of active steroids accessible to receptors and available for metabolism. Differences in the expression of FMO1, NAT12, HSD17B2 and HSD17B13 were verified by quantitative real competitive PCR.

Conclusion

A number of genes and pathways related to metabolism of androstenone in liver were identified, including new candidate genes involved in phase I oxidation metabolism, phase II conjugation metabolism, and regulation of steroid availability. The study is a first step towards a deeper understanding of enzymes and regulators involved in pathways of androstenone metabolism and may ultimately lead to the discovery of markers to reduce boar taint.

     

Integrated analysis of leaf morphological and color traits in different populations of Chinese cabbage (<Emphasis Type="Italic">Brassica rapa</Emphasis> ssp. <Emphasis Type="Italic">pekinensis</Emphasis>)

Key message

QTLs and candidate gene markers associated with leaf morphological and color traits were identified in two immortalized populations of Brassica rapa, which will provide genetic information for marker-assisted breeding.

Abstract

Brassica rapa is an important leafy vegetable consumed worldwide and morphology is a key character for its breeding. To enhance genetic control, quantitative trait loci (QTLs) for leaf color and plant architecture were identified using two immortalized populations with replications of 2 and 4 years. Overall, 158 and 80 QTLs associated with 23 and 14 traits were detected in the DH and RIL populations, respectively. Among them, 23 common robust-QTLs belonging to 12 traits were detected in common loci over the replications. Through comparative analysis, five crucifer genetic blocks corresponding to morphology trait (R, J&U, F and E) and color trait (F, E) were identified in three major linkage groups (A2, A3 and A7). These might be key conserved genomic regions involved with the respective traits. Through synteny analysis with Arabidopsis, 64 candidate genes involved in chlorophyll biosynthesis, cell proliferation and elongation were co-localized within QTL intervals. Among them, SCO3, ABI3, FLU, HCF153, HEMB1, CAB3 were mapped within QTLs for leaf color; and CYCD3;1, CYCB2;4, AN3, ULT1 and ANT were co-localized in QTL regions for leaf size. These robust QTLs and their candidate genes provide useful information for further research into leaf architecture with crop breeding.