Efficient Mixed-model Association Research Articles

Genome-wide association study identified five quantitative trait loci and two candidate genes for digestive traits in Suhuai pigs.

This work aimed to identify markers and candidate genes underlying porcine digestive traits. In total, 331 pigs were genotyped by 80 K Chip data or 50 K Chip data. For apparent neutral detergent fiber digestibility, a total of 19 and 21 candidate single nucleotide polymorphisms (SNP) were respectively identified using a genome-wide efficient mixed-model association algorithm and linkage-disequilibrium adjusted kinship. Among them, three quantitative trait locus (QTL) regions were identified. For apparent acid detergent fiber digestibility, a total of 16 and 17 SNPs were identified by these two methods, respectively. Of these, three QTL regions were also identified. Moreover, two candidate genes (MST1 and LATS1), which are functionally related to intestinal homeostasis and health, were detected near these significant SNPs. Taken together, our results could provide a basis for deeper research on digestive traits in pigs.

Identification of Single Nucleotide Polymorphic Loci and Candidate Genes for Seed Germination Percentage in Okra under Salt and No-Salt Stresses by Genome-Wide Association Study.

Excessive soil salinity is a major stressor inhibiting crops' growth, development, and yield. Seed germination is a critical stage of crop growth and development, as well as one of the most salt-sensitive stages. Salt stress has a significant inhibitory effect on seed germination. Okra is a nutritious vegetable, but its seed germination percentage (GP) is low, whether under salt stress conditions or suitable conditions. In this study, we used 180 okra accessions and conducted a genome-wide association study (GWAS) on the germination percentage using 20,133,859 single nucleotide polymorphic (SNP) markers under 0 (CK, diluted water), 70 (treatment 1, T1), and 140 mmol/L (treatment 2, T2) NaCl conditions. Using the mixed linear model (MLM) in Efficient Mixed-model Association eXpedated (EMMAX) and Genome-wide Efficient Mixed Model Association (GEMMA) software, 511 SNP loci were significantly associated during germination, of which 167 SNP loci were detected simultaneously by both programs. Among the 167 SNPs, SNP2619493 on chromosome 59 and SNP2692266 on chromosome 44 were detected simultaneously under the CK, T1, and T2 conditions, and were key SNP loci regulating the GP of okra seeds. Linkage disequilibrium block analysis revealed that nsSNP2626294 (C/T) in Ae59G004900 was near SNP2619493, and the amino acid changes caused by nsSNP2626294 led to an increase in the phenotypic values in some okra accessions. There was an nsSNP2688406 (A/G) in Ae44G005470 near SNP2692266, and the amino acid change caused by nsSNP2688406 led to a decrease in phenotypic values in some okra accessions. These results indicate that Ae59G004900 and Ae44G005470 regulate the GP of okra seeds under salt and no-salt stresses. The gene expression analysis further demonstrated these results. The SNP markers and genes that were identified in this study will provide reference for further research on the GP of okra, as well as new genetic markers and candidate genes for cultivating new okra varieties with high GPs under salt and no-salt stress conditions.

A metabolomics study in citrus provides insight into bioactive phenylpropanoid metabolism.

Abstract Citrus fruits are widely consumed worldwide in juices or as fresh and provide a broad range of phytonutrients that are important for human health. Here, a citrus multi-omics resource is presented: comprehensive metabolic profiling of various citrus species was performed and metabolic profiles were compared among species, with a focus on the phenylpropanoid metabolic pathway. A metabolite-based genome-wide association analysis (mGWAS) of 154 pummelo accessions was performed using factored spectrally transformed linear mixed models (FaST-LMM) and efficient mixed-model association eXpedited (EMMAX), and the genetic and biochemical basis of metabolomic variation was comprehensively analysed. A metabolite-single nucleotide polymorphism-gene (metabolite-SNP-gene) interaction network was constructed for pummelo, and many candidate loci controlling the synthesis and regulation of bioactive compounds were identified; among these loci, three BAHD malonyltransferases were involved in the malonylation of flavonoid glycosides. Further investigation revealed that an R2R3-MYB transcription factor CgMYB1 positively controls the metabolism of phenylpropanoid molecules, particularly the flavonoid derivatives. This study provides valuable data resources on the metabolic regulatory networks of bioactive components in citrus, in addition to demonstrating an efficient method for metabolic pathway dissection and providing targets for future breeding work with the aim of improving nutritional value.

Genome-wide association study of powdery mildew resistance in cultivated soybean from Northeast China.

Powdery mildew (PMD), caused by the pathogen Microsphaera diffusa, leads to substantial yield decreases in susceptible soybean under favorable environmental conditions. Effective prevention of soybean PMD damage can be achieved by identifying resistance genes and developing resistant cultivars. In this study, we genotyped 331 soybean germplasm accessions, primarily from Northeast China, using the SoySNP50K BeadChip, and evaluated their resistance to PMD in a greenhouse setting. To identify marker-trait associations while effectively controlling for population structure, we conducted genome-wide association studies utilizing factored spectrally transformed linear mixed models, mixed linear models, efficient mixed-model association eXpedited, and compressed mixed linear models. The results revealed seven single nucleotide polymorphism (SNP) loci strongly associated with PMD resistance in soybean. Among these, one SNP was localized on chromosome (Chr) 14, and six SNPs with low linkage disequilibrium were localized near or in the region of previously mapped genes on Chr 16. In the reference genome of Williams82, we discovered 96 genes within the candidate region, including 17 resistance (R)-like genes, which were identified as potential candidate genes for PMD resistance. In addition, we performed quantitative real-time reverse transcription polymerase chain reaction analysis to evaluate the gene expression levels in highly resistant and susceptible genotypes, focusing on leaf tissues collected at different times after M. diffusa inoculation. Among the examined genes, three R-like genes, including Glyma.16G210800, Glyma.16G212300, and Glyma.16G213900, were identified as strong candidates associated with PMD resistance. This discovery can significantly enhance our understanding of soybean resistance to PMD. Furthermore, the significant SNPs strongly associated with resistance can serve as valuable markers for genetic improvement in breeding M. diffusa-resistant soybean cultivars.

Genome-wide association studies to identify quantitative trait loci and positional candidate genes affecting meat quality-related traits in pigs.

Meat quality comprises a set of key traits such as pH, meat color, water-holding capacity, tenderness and marbling. These traits are complex because they are affected by multiple genetic and environmental factors. The aim of this study was to investigate the molecular genetic basis underlying nine meat quality-related traits in a Yorkshire pig population using a genome-wide association study (GWAS) and subsequent biological pathway analysis. In total, 45,926 single nucleotide polymorphism (SNP) markers from 543 pigs were selected for the GWAS after quality control. Data were analyzed using a genome-wide efficient mixed model association (GEMMA) method. This linear mixed model-based approach identified two quantitative trait loci (QTLs) for meat color (b*) on chromosome 2 (SSC2) and one QTL for shear force on chromosome 8 (SSC8). These QTLs acted additively on the two phenotypes and explained 3.92%-4.57% of the phenotypic variance of the traits of interest. The genes encoding HAUS8 on SSC2 and an lncRNA on SSC8 were identified as positional candidate genes for these QTLs. The results of the biological pathway analysis revealed that positional candidate genes for meat color (b*) were enriched in pathways related to muscle development, muscle growth, intramuscular adipocyte differentiation, and lipid accumulation in muscle, whereas positional candidate genes for shear force were overrepresented in pathways related to cell growth, cell differentiation, and fatty acids synthesis. Further verification of these identified SNPs and genes in other independent populations could provide valuable information for understanding the variations in pork quality-related traits.

Genome-wide association study of feed conversion ratio in turbot (Scophthalmus maximus) based on genome resequencing

The feed conversion rate (FCR) is a crucial economic trait in aquaculture fish, as feed costs account for the vast majority of aquaculture costs. Given the importance of this trait, there have little efforts to unravel the genomic architecture of FCR and identify key genes or genomic regions useful as selection makers. This situation may stem from the inherent swimming behavior of fish, which presents challenges in obtaining individual FCRs. In this work, we developed a specialized small cage culture system. Through this system, the Feed Conversion Ratio (FCR) of 300 individuals was successfully determined, marking the first successful determination of individual FCRs for turbot. The results revealed a range of FCR values from 35.3% to 192.6%, with an average of 126.5% ± 23.5%. On this basis, a genome-wide association study (GWAS) was conducted to identify single nucleotide polymorphisms (SNP) associated with FCR. A total of 2613,115 SNPs on 22 chromosomes were selected for GWAS analysis by Efficient Mixed-Model Association eXpedited (EMMAX) software. A linear mixed model was employed to test the association between each SNP and FCR, considering population structure and genetic relatedness among individuals. Two significant SNPs on Chr6 were identified to be correlated with FCR. According to their localization in genome, four candidate genes were identified on Chr6, including uncharacterized protein KIAA1755, Kinesin Family Member 21B (KIF21B), Discs large homolog-associated protein 4B (DLGAP4B), and Copine 5B (CPNE5B), all of which are closely located in the genome and exhibit significant functional similarities. Our results will facilitate the implementation of marker-assisted selection in breeding programs and serve as a foundation for future research in this area.

Genome-wide association study reveals genomic loci of sex differentiation and gonadal development in Plectropomus leopardus.

Introduction: Plectropomus leopardus, a commercially significant marine fish, is primarily found in the Western Pacific regions and along the coast of Southeast Asia. A thorough analysis of the molecular mechanisms involved in sex differentiation is crucial for gaining a comprehensive understanding of gonadal development and improving sex control breeding. However, the relevant fundamental studies of P. leopardus are relatively lacking. Methods: In this study, a genome-wide association study (GWAS) was conducted to investigate the genetic basis mechanism of sex differentiation and gonadal developmental traits in P. leopardus utilizing about 6,850,000 high-quality single-nucleotide polymorphisms (SNPs) derived from 168 individuals (including 126 females and 42 males) by the genome-wide efficient mixed-model association (GEMMA) algorithm. Results: The results of these single-trait GWASs showed that 46 SNP loci (-log10 p > 7) significantly associated with sex differentiation, and gonadal development traits were distributed in multiple different chromosomes, which suggested the analyzed traits were all complex traits under multi-locus control. A total of 1,838 potential candidate genes were obtained by considering a less-stringent threshold (-log10 p > 6) and ±100kb regions surrounding the significant genomic loci. Moreover, 31 candidate genes were identified through a comprehensive analysis of significant GWAS peaks, gene ontology (GO) annotations, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses, including taf7, ddx6, apoeb, sgk1, a2m, usf1, hsd3b7, dll4, xbp1, tet3, esr1, and gli3. These trait-associated genes have been shown to be involved in germline development, male sex differentiation, gonad morphogenesis, hormone receptor binding, oocyte development, male gonad development, steroidogenesis, estrogen-synthetic pathway, etc. Discussion: In the present study, multiple genomic loci of P. leopardus associated with sex differentiation and gonadal development traits were identified for the first time by using GWAS, providing a valuable resource for further research on the molecular genetic mechanism and sex control in P. leopardus. Our results also can contribute to understanding the genetic basis of the sex differentiation mechanism and gonadal development process in grouper fish.

Genome-wide association study using specific-locus amplified fragment sequencing identifies new genes influencing nitrogen use efficiency in rice landraces.

Nitrogen is essential for crop production. It is a critical macronutrient for plant growth and development. However, excessive application of nitrogen fertilizer is not only a waste of resources but also pollutes the environment. An effective approach to solving this problem is to breed rice varieties with high nitrogen use efficiency (NUE). In this study, we performed a genome-wide association study (GWAS) on 419 rice landraces using 208,993 single nucleotide polymorphisms (SNPs). With the mixed linear model (MLM) in the Tassel software, we identified 834 SNPs associated with root surface area (RSA), root length (RL), root branch number (RBN), root number (RN), plant dry weight (PDW), plant height (PH), root volume (RL), plant fresh weight (PFW), root fractal dimension (RFD), number of root nodes (NRN), and average root diameter (ARD), with a significant level of p < 2.39×10-7. In addition, we found 49 SNPs that were correlated with RL, RBN, RN, PDW, PH, PFW, RFD, and NRN using genome-wide efficient mixed-model association (GEMMA), with a significant level of p < 1×10-6. Additionally, the final results for eight traits associated with 193 significant SNPs by using multi-locus random-SNP-effect mixed linear model (mrMLM) model and 272 significant SNPs associated with 11 traits by using IIIVmrMLM. Within the linkage intervals of significantly associated SNP, we identified eight known related genes to NUE in rice, namely, OsAMT2;3, OsGS1, OsNR2, OsNPF7.4, OsPTR9, OsNRT1.1B, OsNRT2.3, and OsNRT2.2. According to the linkage disequilibrium (LD) decay value of this population, there were 75 candidate genes within the 150-kb regions upstream and downstream of the most significantly associated SNP (Chr5_29804690, Chr5_29956584, and Chr10_17540654). These candidate genes included 22 transposon genes, 25 expressed genes, and 28 putative functional genes. The expression levels of these candidate genes were measured by real-time quantitative PCR (RT-qPCR), and the expression levels of LOC_Os05g51700 and LOC_Os05g51710 in C347 were significantly lower than that in C117; the expression levels of LOC_Os05g51740, LOC_Os05g51780, LOC_Os05g51960, LOC_Os05g51970, and LOC_Os10g33210 were significantly higher in C347 than C117. Among them, LOC_Os10g33210 encodes a peptide transporter, and LOC_Os05g51690 encodes a CCT domain protein and responds to NUE in rice. This study identified new loci related to NUE in rice, providing new genetic resources for the molecular breeding of rice landraces with high NUE.

Estimation of genetic parameters for pork belly traits.

Pork belly is a cut of meat with high worldwide demand. However, although the belly is comprised of multiple muscles and fat, unlike the loin muscle, research on their genetic parameters has yet to focus on a representative cut. To use swine breeding, it is necessary to estimate heritability against pork belly traits. Moreover, estimating genetic correlations is needed to identify genetic relationship among the traditional carcass and meat quality traits. This study sought to estimate the heritability of the carcass, belly, and their component traits, as well as the genetic correlations among them, to confirm whether these traits can be improved. A total of 543 Yorkshire pigs (406 castrated males and 137 females) from 49 sires and 244 dam were used in this study. To estimate genetic parameters, a total of 12 traits such as lean meat production ability, meat quality and pork belly traits were chosen. The heritabilities were estimated by using genome-wide efficient mixed model association software. The statistical model was selected so that farm, carcass weight, sex, and slaughter season were fixed effects. In addition, its genetic parameters were calculated via MTG2 software. The heritability estimates for the 7th belly slice along the whole plate and its components were low to moderate (0.07±0.07 to 0.33±0.07). Moreover, the genetic correlations among the carcass and belly traits were moderate to high (0.28±0.20 to 0.99±0.31). Particularly, the rectus abdominis muscle exhibited a high absolute genetic correlation with the belly and meat quality (0.73±52 to 0.93±0.43). A moderate to high correlation coefficient was obtained based on the genetic parameters. The belly could be genetically improved to contain a larger proportion of muscle regardless of lean meat production ability.

GWAS resolves molecular mechanisms underlying natural variation for carotenoids in Cucurbita maxima Duchesne

Carotenoid levels and composition are essential nutraceutical traits in Cucurbita maxima influencing the fruit flesh and rind colors. In this study, Mr Fugly, Boston Marrow, Baby Green Hubbard, Chicago Warted Hubbard, Golden Hubbard, and Bush Buttercup of USA; Australian Butter (Australia); Marmellata and Marina di Chioggia of Italy contained the highest carotenoid content. EMMAX (Efficient Mixed-Model Association eXpedited) was employed for a Genome-wide association study (GWAS) to identify associated genomic regions underlying the regulation of carotenoids. Here, we profiled the contents of α-carotene, β-carotene, β-cryptoxanthin, lutein, and zeaxanthin from a diversity panel of C. maxima consisting of classical horticulture groups including buttercup, turban, banana, zapallito, hubbard, mammoth and australian. Lutein and β-carotene were the major carotenoids among C. maxima accessions. Furthermore, we used genotyping by sequencing to identify 12,996 single nucleotide polymorphisms (SNPs) that were used to resolve population structure with three primary clusters and their admixed genotypes. Our GWAS revealed several SNPs related to 289 genes involved in plastid biogenesis, protein ubiquitination, transcriptional regulation at the carotenoid biosynthesis level, fruit ripening, and regulation of carotenoid transport. Together, these results contribute to a better understanding of the molecular mechanisms of carotenoid biosynthesis in fruit flesh and would facilitate breeding for enhanced carotenoids in winter squashes.

Genetic Dissection of Grain Size Traits Through Genome-Wide Association Study Based on Genic Markers in Rice

Grain size plays a significant role in rice, starting from affecting yield to consumer preference, which is the driving force for deep investigation and improvement of grain size characters. Quantitative inheritance makes these traits complex to breed on account of several alleles contributing to the complete trait expression. We employed genome-wide association study in an association panel of 88 rice genotypes using 142 new candidate gene based SSR (cgSSR) markers, derived from yield-related candidate genes, with the efficient mixed-model association coupled mixed linear model for dissecting complete genetic control of grain size traits. A total of 10 significant associations were identified for four grain size-related characters (grain weight, grain length, grain width, and length-width ratio). Among the identified associations, seven marker trait associations explain more than 10% of the phenotypic variation, indicating major putative QTLs for respective traits. The allelic variations at genes OsBC1L4 , SHO1 and OsD2 showed association between 1000-grain weight and grain width, 1000-grain weight and grain length, and grain width and length-width ratio, respectively. The cgSSR markers, associated with corresponding traits, can be utilized for direct allelic selection, while other significantly associated cgSSRs may be utilized for allelic accumulation in the breeding programs or grain size improvement. The new cgSSR markers associated with grain size related characters have a significant impact on practical plant breeding to increase the number of causative alleles for these traits through marker aided rice breeding programs.

Genome-Wide Association Study and Selective Sweep Analysis Reveal the Genetic Architecture of Body Weights in a Chicken F2 Resource Population.

Rapid growth is one of the most important economic traits in broiler breeding programs. Identifying markers and genes for growth traits may not only benefit marker-assisted selection (MAS)/genomic selection (GS) but also provide important information for understanding the genetic architecture of growth traits in broilers. In the present study, an F2 resource population derived from a cross between the broiler and Baier yellow chicken (a Chinese local breed) was used and body weights from 1 to 12 weeks of age [body weight (BW) 1–BW12)] were measured. A total of 519 F2 birds were genome re-sequenced, and a combination of genome-wide association study (GWAS) and selective sweep analysis was carried out to characterize the genetic architecture affecting chicken body weight comprehensively. As a result, 1,539 SNPs with significant effects on body weights at different weeks of age were identified using a genome-wide efficient mixed-model association (GEMMA) package. These SNPs were distributed on chromosomes 1 and 4. Besides, windows under selection identified for BW1–BW12 varied from 1,581 to 2,265. A total of 42 genes were also identified with significant effects on BW1–BW12 based on both GWAS and selective sweep analysis. Among these genes, diacylglycerol kinase eta (DGKH), deleted in lymphocytic leukemia (DLEU7), forkhead box O17 (FOXO1), karyopherin subunit alpha 3 (KPNA3), calcium binding protein 39 like (CAB39L), potassium voltage-gated channel interacting protein 4 (KCNIP4), and slit guidance ligand 2 (SLIT2) were considered as important genes for broiler growth based on their basic functions. The results of this study may supply important information for understanding the genetic architecture of growth traits in broilers.

Genetics of Base Coat Colour Variations and Coat Colour-Patterns of the South African Nguni Cattle Investigated Using High-Density SNP Genotypes.

Nguni cattle are a Sanga type breed with mixed B. taurus and B. indicus ancestry and proven resistance to ticks, diseases and other harsh conditions of the African geographical landscape. The multi-coloured Nguni coats have found a niche market in the leather industry leading to breeding objectives towards the promotion of such diversity. However, there is limited studies on the genomic architecture underlying the coat colour and patterns hampering any potential breeding and improvement of such trait. This study investigated the genetics of base coat colour, colour-sidedness and the white forehead stripe in Nguni cattle using coat colour phenotyped Nguni cattle and Illumina Bovine HD (770K) genotypes. Base coat colour phenotypes were categorised into eumelanin (n = 45) and pheomelanin (n = 19). Animals were categorised into either colour-sided (n = 46) or non-colour-sided (n = 94) and similarly into presence (n = 15) or absence (n = 67) of white forehead stripe. Genome-wide association tests were conducted using 622,103 quality controlled SNPs and the Efficient Mixed Model Association eXpedited method (EMMAX) implemented in Golden Helix SNP Variation Suite. The genome-wide association studies for base coat colour (eumelanin vs. pheomelanin) resulted into four indicative SNPs on BTA18 and a well-known gene, MC1R, was observed within 1 MB from the indicative SNPs (p < 0.00001) and found to play a role in the melanogenesis (core pathway for melanin production) and the MAPK signalling pathway. GWAS for colour-sidedness resulted in four indicative SNPs, none of which were in close proximity to the KIT candidate gene known for colour-sidedness. GWAS for the white forehead stripe resulted in 17 indicative SNPs on BTA6. Four genes MAPK10, EFNA5, PPP2R3C and PAK1 were found to be associated with the white forehead stripe and were part of the MAPK, adrenergic and Wnt signalling pathways that are synergistically associated with the synthesis of melanin. Overall, our results prove prior knowledge of the role of MC1R in base coat colours in cattle and suggested a different genetic mechanism for forehead stripe phenotypes in Nguni cattle.

New Insights on Gene by Environmental Effects of Drugs of Abuse in Animal Models Using GeneNetwork.

Gene-by-environment interactions are important for all facets of biology, especially behaviour. Families of isogenic strains of mice, such as the BXD strains, are excellently placed to study these interactions, as the same genome can be tested in multiple environments. BXD strains are recombinant inbred mouse strains derived from crossing two inbred strains—C57BL/6J and DBA/2J mice. Many reproducible genometypes can be leveraged, and old data can be reanalysed with new tools to produce novel insights. We obtained drug and behavioural phenotypes from Philip et al. Genes, Brain and Behaviour 2010, and reanalysed their data with new genotypes from sequencing, as well as new models (Genome-wide Efficient Mixed Model Association (GEMMA) and R/qtl2). We discovered QTLs on chromosomes 3, 5, 9, 11, and 14, not found in the original study. We reduced the candidate genes based on their ability to alter gene expression or protein function. Candidate genes included Slitrk6 and Cdk14. Slitrk6, in a Chromosome14 QTL for locomotion, was found to be part of a co-expression network involved in voluntary movement and associated with neuropsychiatric phenotypes. Cdk14, one of only three genes in a Chromosome5 QTL, is associated with handling induced convulsions after ethanol treatment, that is regulated by the anticonvulsant drug valproic acid. By using families of isogenic strains, we can reanalyse data to discover novel candidate genes involved in response to drugs of abuse.

An Efficient Score Test Integrated with Empirical Bayes for Genome-Wide Association Studies.

Many methods used in multi-locus genome-wide association studies (GWAS) have been developed to improve statistical power. However, most existing multi-locus methods are not quicker than single-locus methods. To address this concern, we proposed a fast score test integrated with Empirical Bayes (ScoreEB) for multi-locus GWAS. Firstly, a score test was conducted for each single nucleotide polymorphism (SNP) under a linear mixed model (LMM) framework, taking into account the genetic relatedness and population structure. Then, all of the potentially associated SNPs were selected with a less stringent criterion. Finally, Empirical Bayes in a multi-locus model was performed for all of the selected SNPs to identify the true quantitative trait nucleotide (QTN). Our new method ScoreEB adopts the similar strategy of multi-locus random-SNP-effect mixed linear model (mrMLM) and fast multi-locus random-SNP-effect EMMA (FASTmrEMMA), and the only difference is that we use the score test to select all the potentially associated markers. Monte Carlo simulation studies demonstrate that ScoreEB significantly improved the computational efficiency compared with the popular methods mrMLM, FASTmrEMMA, iterative modified-sure independence screening EM-Bayesian lasso (ISIS EM-BLASSO), hybrid of restricted and penalized maximum likelihood (HRePML) and genome-wide efficient mixed model association (GEMMA). In addition, ScoreEB remained accurate in QTN effect estimation and effectively controlled false positive rate. Subsequently, ScoreEB was applied to re-analyze quantitative traits in plants and animals. The results show that ScoreEB not only can detect previously reported genes, but also can mine new genes.

Genome-wide association study identifies genomic loci of sex determination, gonadal weight and gonadosomatic index traits in Takifugu bimaculatus

Takifugu bimaculatus is a cultured marine teleost fish, distributed ranging from the southern Yellow Sea to the South China Sea. Understanding the mechanism of sex determination is essential for sex control breeding biotechnologies of T. bimaculatus. However, there lacks a rapid and accurate sex specific marker for the T. bimaculatus. In this study, whole genome resequencing was conducted on 71 individuals (including 49 females and 22 males), 331,190 high-quality polymorphic loci (34,435 INDEL, 296,755 single-nucleotide polymorphisms (SNPs)) were obtained through genome-wide association analysis (GWAS) by GEMMA (Genome-wide Efficient Mixed Model Association algorithm). 1403 loci on chromosome 7 were identified as sex significantly associated loci (P < 1.51 × 10−7), and a 12 Mb sex-associated region on Chr7 was narrowed down by Fst scanning. It was worth noting that the SNP, SNP1100879 on Chr7 at position 8,764,381 was identified to be sex determination in T. bimaculatus. 13 pairs of primers for sex identification Marker-F/R were designed and it was validated by PCR. However, only one pair of primers (SNP-7F/R) was sex-linked, in which 95% of female genotypes are homozygous CC and 88% of male genotypes are heterozygous CT, suggesting a XX/XY sex determination system in T. bimaculatus. In addition, 12 sex-related candidate genes were identified around 306 SNPs, including amhr2, dzip1, patz1, brdt, hmga2, gli1, inhbb, mmp19, wnt2b, taf4 and so on. Because of the significant difference of gonadal weight (GW) and gonadosomatic index (GSI) between females and males of T. bimaculatus, GWAS on GW and GSI of different genders were also conducted independently. Besides, we identified 11 SNPs, located on Chr7, Chr18, Chr21, and Chr2, showing genome-wide significant association with GW and GSI and five genes including cyp27b1, hexb, inhba, cgba and shbg, involved in oogenesis, ovarian follicle development, and regulation of estradiol secretion. Then, we performed the gene ontology (GO) and KEGG signaling pathway (KEGG) and Protein-protein interaction (PPI) network analyses of the candidate genes. amhr2, gli1, ihbb and dzip1 genes were identified as hub genes. In the present study, a sex-linked marker of T. bimaculatus was identified for the first time using whole-genome resequencing technology, and its limited versatility provided important clues for deciphering the sex determination mechanism in T. bimaculatus.

Genome-wide hierarchical mixed model association analysis.

In genome-wide mixed model association analysis, we stratified the genomic mixed model into two hierarchies to estimate genomic breeding values (GBVs) using the genomic best linear unbiased prediction and statistically infer the association of GBVs with each SNP using the generalized least square. The hierarchical mixed model (Hi-LMM) can correct confounders effectively with polygenic effects as residuals for association tests, preventing potential false-negative errors produced with genome-wide rapid association using mixed model and regression or an efficient mixed-model association expedited (EMMAX). Meanwhile, the Hi-LMM performs the same statistical power as the exact mixed model association and the same computing efficiency as EMMAX. When the GBVs have been estimated precisely, the Hi-LMM can detect more quantitative trait nucleotides (QTNs) than existing methods. Especially under the Hi-LMM framework, joint association analysis can be made straightforward to improve the statistical power of detecting QTNs.

Identifying Loci Associated With Bovine Corona Virus Infection and Bovine Respiratory Disease in Dairy and Feedlot Cattle

Bovine coronavirus (BCoV) is associated with respiratory and enteric infections in both dairy and beef cattle worldwide. It is also one of a complex of pathogens associated with bovine respiratory disease (BRD), which affects millions of cattle annually. The objectives of this study were to identify loci and heritability estimates associated with BCoV infection and BRD in dairy calves and feedlot cattle. Dairy calves from California (n = 1,938) and New Mexico (n = 647) and feedlot cattle from Colorado (n = 915) and Washington (n = 934) were tested for the presence of BCoV when classified as BRD cases or controls following the McGuirk scoring system. Two comparisons associated with BCoV were investigated: (1) cattle positive for BCoV (BCoV+) were compared to cattle negative for BCoV (BCoV−) and (2) cattle positive for BCoV and affected with BRD (BCoV+BRD+) were compared to cattle negative for BCoV and BRD (BCoV−BRD−). The Illumina BovineHD BeadChip was used for genotyping, and genome-wide association analyses (GWAA) were performed using EMMAX (efficient mixed-model association eXpedited). The GWAA for BCoV+ identified 51 loci (p < 1 × 10−5; 24 feedlot, 16 dairy, 11 combined) associated with infection with BCoV. Three loci were associated with BCoV+ across populations. Heritability estimates for BCoV+ were 0.01 for dairy, 0.11 for feedlot cattle, and 0.03 for the combined population. For BCoV+BRD+, 80 loci (p < 1 × 10−5; 26 feedlot, 25 dairy, 29 combined) were associated including 14 loci across populations. Heritability estimates for BCoV+BRD+ were 0.003 for dairy, 0.44 for feedlot cattle, and 0.07 for the combined population. Several positional candidate genes associated with BCoV and BRD in this study have been associated with other coronaviruses and respiratory infections in humans and mice. These results suggest that selection may reduce susceptibility to BCoV infection and BRD in cattle.

Genome-Wide Association Study (GWAS) of dental caries in diverse populations

BackgroundDental caries is one of the most common chronic diseases and is influenced by a complex interplay of genetic and environmental factors. Most previous genetic studies of caries have focused on identifying genes that contribute to dental caries in specific ethnic groups, usually of European descent.MethodsThe aim of this study is to conduct a genome-wide association study (GWAS) to identify associations affecting susceptibility to caries in a large multiethnic population from Argentina, the Philippines, Guatemala, Hungary, and the USA, originally recruited for studies of orofacial clefts (POFC, N = 3686). Ages of the participants ranged from 2 to 12 years for analysis of the primary dentition, and 18–60 years for analysis of the permanent dentition. For each participant, dental caries was assessed by counts of decayed and filled teeth (dft/DFT) and genetic variants (single nucleotide polymorphisms, SNPs) were genotyped or imputed across the entire genome. Caries was analyzed separately for the primary and permanent dentitions, with age, gender, and presence/absence of any type of OFC treated as covariates. Efficient Mixed-Model Association eXpedited (EMMAX) was used to test genetic association, while simultaneously accounting for relatedness and stratification.ResultsWe identified several suggestive loci (5 × 10−8 < P < 5 × 10−6) within or near genes with plausible biological roles for dental caries, including a cluster of taste receptor genes (TAS2R38, TAS2R3, TAS2R4, TASR25) on chromosome 7 for the permanent dentition analysis, and DLX3 and DLX4 on chromosome 17 for the primary dentition analysis. Genome-wide significant results were seen with SNPs in the primary dentition only; however, none of the identified genes near these variants have known roles in cariogenesis.ConclusionThe results of this study warrant further investigation and may lead to a better understanding of cariogenesis in diverse populations, and help to improve dental caries prediction, prevention, and/or treatment in future.

Accounting for Population Structure and Phenotypes From Relatives in Association Mapping for Farm Animals: A Simulation Study.

Population structure or genetic relatedness should be considered in genome association studies to avoid spurious association. The most used methods for genome-wide association studies (GWAS) account for population structure but are limited to genotyped individuals with phenotypes. Single-step GWAS (ssGWAS) can use phenotypes from non-genotyped relatives; however, its ability to account for population structure has not been explored. Here we investigate the equivalence among ssGWAS, efficient mixed-model association expedited (EMMAX), and genomic best linear unbiased prediction GWAS (GBLUP-GWAS), and how they differ from the single-SNP analysis without correction for population structure (SSA-NoCor). We used simulated, structured populations that mimicked fish, beef cattle, and dairy cattle populations with 1040, 5525, and 1,400 genotyped individuals, respectively. Larger populations were also simulated that had up to 10-fold more genotyped animals. The genomes were composed by 29 chromosomes, each harboring one QTN, and the number of simulated SNPs was 35,000 for the fish and 65,000 for the beef and dairy cattle populations. Males and females were genotyped in the fish and beef cattle populations, whereas only males had genotypes in the dairy population. Phenotypes for a trait with heritability varying from 0.25 to 0.35 were available in both sexes for the fish population, but only for females in the beef and dairy cattle populations. In the latter, phenotypes of daughters were projected into genotyped sires (i.e., deregressed proofs) before applying EMMAX and SSA-NoCor. Although SSA-NoCor had the largest number of true positive SNPs among the four methods, the number of false negatives was two–fivefold that of true positives. GBLUP-GWAS and EMMAX had a similar number of true positives, which was slightly smaller than in ssGWAS, although the difference was not significant. Additionally, no significant differences were observed when deregressed proofs were used as pseudo-phenotypes in EMMAX compared to daughter phenotypes in ssGWAS for the dairy cattle population. Single-step GWAS accounts for population structure and is a straightforward method for association analysis when only a fraction of the population is genotyped and/or when phenotypes are available on non-genotyped relatives.