Glutamate Receptor 6 Gene (<Emphasis Type="Italic">GluR6</Emphasis> or <Emphasis Type="Italic">GRIK2</Emphasis>) Polymorphisms in the Indian Population: A Genetic Association Study on Autism Spectrum Disorder

Autism is a neurodevelopmental disorder with early manifestation. It is a multifactorial disorder and several susceptible chromosomal regions for autism are identified through genome scan studies. The gene coding for glutamate receptor 6 (GluR6 or GRIK2) has been suggested as a candidate gene for autism based on its localization in the autism specific region on chromosome 6q21 and the involvement of receptor protein in cognitive functions like learning and memory. Despite its importance, so far no studies have been carried out on possible involvement of GluR6 with autism in the Indian population. Therefore in the present study, we have performed genetic analysis of three markers of GluR6 (SNP1: rs2227281, SNP2: rs2227283, SNP3: rs2235076) for possible association with autism through population, and family-based (TDT and HHRR) approaches. DSM-IV criteria and CARS/ADI-R have been utilized for diagnosis. Genotyping analysis for the SNPs has been carried out in 101 probands with autism spectrum disorder, 180 parents and 152 controls from different regions of India. Since the minor allele frequency of SNP3 was too low, the association studies have been carried out only for SNP1 and SNP2. Even though two earlier studies have shown association of these markers with autism, the present case–control and TDT, as well as HHRR analyses have not demonstrated any biased transmission of alleles or haplotypes to the affected offspring. Thus our results suggest that these markers of GluR6 are unlikely to be associated with autism in the Indian population.     

Genotyping‐by‐sequencing‐based genome‐wide association studies on Verticillium wilt resistance in autotetraploid alfalfa (Medicago sativa L.)

Verticillium wilt (VW) is a fungal disease that causes severe yield losses in alfalfa. The most effective method to control the disease is through the development and use of resistant varieties. The identification of marker loci linked to VW resistance can facilitate breeding for disease‐resistant alfalfa. In the present investigation, we applied an integrated framework of genome‐wide association with genotyping‐by‐sequencing (GBS) to identify VW resistance loci in a panel of elite alfalfa breeding lines. Phenotyping was performed by manual inoculation of the pathogen to healthy seedlings, and scoring for disease resistance was carried out according to the standard test of the North America Alfalfa Improvement Conference (NAAIC). Marker–trait association by linkage disequilibrium identified 10 single nucleotide polymorphism (SNP) markers significantly associated with VW resistance. Alignment of the SNP marker sequences to the M. truncatula genome revealed multiple quantitative trait loci (QTLs). Three, two, one and five markers were located on chromosomes 5, 6, 7 and 8, respectively. Resistance loci found on chromosomes 7 and 8 in the present study co‐localized with the QTLs reported previously. A pairwise alignment (blastn ) using the flanking sequences of the resistance loci against the M. truncatula genome identified potential candidate genes with putative disease resistance function. With further investigation, these markers may be implemented into breeding programmes using marker‐assisted selection, ultimately leading to improved VW resistance in alfalfa.     

Genome‐wide association study of growth traits in Jinghai Yellow chicken hens using SLAF‐seq technology

In this study, we performed a new genome‐wide association study using SLAF‐seq technology. A total of 19 single nucleotide polymorphism effects involving nine different SNP markers reached 5% Bonferroni‐corrected genome‐wide significance. In addition, a 5‐Mb region spanning 72.9–77.9 Mb on GGA4, exhibiting many significant SNP effects, was identified. The LDB2 gene in this region had a very strong association with body weight. Another SNP on GGA1, located in the INTS6 gene, had the strongest association with late body weight (weeks 10–16). Some of the SNPs that reached suggestive significance level overlapped with previously reported quantitative trait locus regions.     

Genome‐wide association analysis identifies quantitative trait loci for growth in a Landrace purebred population

Growth‐related traits are complex and economically important in the livestock industry. The aim of this study was to identify quantitative trait loci (QTL) and the associated positional candidate genes affecting growth in pigs. A genome‐wide association study (GWAS) was performed using the porcine single‐nucleotide polymorphism (SNP) 60K bead chip. A mixed‐effects model and linear regression approach were used for the GWAS. The data used in the study included 490 purebred Landrace pigs. All experimental animals were genotyped with 39 438 SNPs located throughout the pig autosomes. We identified a strong association between a SNP marker on chromosome 16 and body weight at 71 days of age (ALGA0092396, P = 5.35 × 10^?9, Bonferroni adjusted P < 0.05). The SNP marker was located near the genomic region containing IRX4, which encodes iroquois homeobox 4. This SNP marker could be useful in the selective breeding program after validating its effect on other populations.     

Haplotype‐based genotyping‐by‐sequencing in oat genome research

In a de novo genotyping‐by‐sequencing (GBS) analysis of short, 64‐base tag‐level haplotypes in 4657 accessions of cultivated oat, we discovered 164741 tag‐level (TL) genetic variants containing 241224 SNPs. From this, the marker density of an oat consensus map was increased by the addition of more than 70000 loci. The mapped TL genotypes of a 635‐line diversity panel were used to infer chromosome‐level (CL) haplotype maps. These maps revealed differences in the number and size of haplotype blocks, as well as differences in haplotype diversity between chromosomes and subsets of the diversity panel. We then explored potential benefits of SNP vs. TL vs. CL GBS variants for mapping, high‐resolution genome analysis and genomic selection in oats. A combined genome‐wide association study (GWAS) of heading date from multiple locations using both TL haplotypes and individual SNP markers identified 184 significant associations. A comparative GWAS using TL haplotypes, CL haplotype blocks and their combinations demonstrated the superiority of using TL haplotype markers. Using a principal component‐based genome‐wide scan, genomic regions containing signatures of selection were identified. These regions may contain genes that are responsible for the local adaptation of oats to Northern American conditions. Genomic selection for heading date using TL haplotypes or SNP markers gave comparable and promising prediction accuracies of up to r = 0.74. Genomic selection carried out in an independent calibration and test population for heading date gave promising prediction accuracies that ranged between r = 0.42 and 0.67. In conclusion, TL haplotype GBS‐derived markers facilitate genome analysis and genomic selection in oat.     

Genome‐wide association study of word reading: Overlap with risk genes for neurodevelopmental disorders

Reading disabilities (RD) are the most common neurocognitive disorder, affecting 5% to 17% of children in North America. These children often have comorbid neurodevelopmental/psychiatric disorders, such as attention deficit/hyperactivity disorder (ADHD). The genetics of RD and their overlap with other disorders is incompletely understood. To contribute to this, we performed a genome‐wide association study (GWAS) for word reading. Then, using summary statistics from neurodevelopmental/psychiatric disorders, we computed polygenic risk scores (PRS) and used them to predict reading ability in our samples. This enabled us to test the shared aetiology between RD and other disorders. The GWAS consisted of 5.3 million single nucleotide polymorphisms (SNPs) and two samples; a family‐based sample recruited for reading difficulties in Toronto (n = 624) and a population‐based sample recruited in Philadelphia [Philadelphia Neurodevelopmental Cohort (PNC)] (n = 4430). The Toronto sample SNP‐based analysis identified suggestive SNPs (P ~ 5 × 10^?7) in the ARHGAP23 gene, which is implicated in neuronal migration/axon pathfinding. The PNC gene‐based analysis identified significant associations (P < 2.72 × 10^?6) for LINC00935 and CCNT1, located in the region of the KANSL2/CCNT1/LINC00935/SNORA2B/SNORA34/MIR4701/ADCY6 genes on chromosome 12q, with near significant SNP‐based analysis. PRS identified significant overlap between word reading and intelligence (R² = 0.18, P = 7.25 × 10^?181), word reading and educational attainment (R² = 0.07, P = 4.91 × 10^?48) and word reading and ADHD (R² = 0.02, P = 8.70 × 10^?6; threshold for significance = 7.14 × 10^?3). Overlap was also found between RD and autism spectrum disorder (ASD) as top‐ranked genes were previously implicated in autism by rare and copy number variant analyses. These findings support shared risk between word reading, cognitive measures, educational outcomes and neurodevelopmental disorders, including ASD.     

Variants in the 1q21 risk region are associated with a visual endophenotype of autism and schizophrenia

Deficits in sensitivity to visual stimuli of low spatial frequency and high temporal frequency (so‐called frequency‐doubled gratings) have been demonstrated both in schizophrenia and in autism spectrum disorder (ASD). Such basic perceptual functions are ideal candidates for molecular genetic study, because the underlying neural mechanisms are well characterized; but they have sometimes been overlooked in favor of cognitive and neurophysiological endophenotypes, for which neural substrates are often unknown. Here, we report a genome‐wide association study of a basic visual endophenotype associated with psychological disorder. Sensitivity to frequency‐doubled gratings was measured in 1060 healthy young adults, and analyzed for association with genotype using linear regression at 642 758 single nucleotide polymorphism (SNP) markers. A significant association (P = 7.9 × 10^?9) was found with the SNP marker rs1797052, situated in the 5′‐untranslated region of PDZK1; each additional copy of the minor allele was associated with an increase in sensitivity equivalent to more than half a standard deviation. A permutation procedure, which accounts for multiple testing, showed that the association was significant at the α = 0.005 level. The region on chromosome 1q21.1 surrounding PDZK1 is an established susceptibility locus both for schizophrenia and for ASD, mirroring the common association of the visual endophenotype with the two disorders. PDZK1 interacts with N‐methyl‐d ‐aspartate receptors and neuroligins, which have been implicated in the etiologies of schizophrenia and ASD. These findings suggest that perceptual abnormalities observed in two different disorders may be linked by common genetic elements .     

Genome‐wide association analyses of child genotype effects and parent‐of‐origin effects in specific language impairment

Specific language impairment (SLI) is a neurodevelopmental disorder that affects linguistic abilities when development is otherwise normal. We report the results of a genome‐wide association study of SLI which included parent‐of‐origin effects and child genotype effects and used 278 families of language‐impaired children. The child genotype effects analysis did not identify significant associations. We found genome‐wide significant paternal parent‐of‐origin effects on chromosome 14q12 (P = 3.74 × 10^?8) and suggestive maternal parent‐of‐origin effects on chromosome 5p13 (P = 1.16 × 10^?7). A subsequent targeted association of six single‐nucleotide‐polymorphisms (SNPs) on chromosome 5 in 313 language‐impaired individuals and their mothers from the ALSPAC cohort replicated the maternal effects, albeit in the opposite direction (P = 0.001); as fathers' genotypes were not available in the ALSPAC study, the replication analysis did not include paternal parent‐of‐origin effects. The paternally‐associated SNP on chromosome 14 yields a non‐synonymous coding change within the NOP9 gene. This gene encodes an RNA‐binding protein that has been reported to be significantly dysregulated in individuals with schizophrenia. The region of maternal association on chromosome 5 falls between the PTGER4 and DAB2 genes, in a region previously implicated in autism and ADHD. The top SNP in this association locus is a potential expression QTL of ARHGEF19 (also called WGEF) on chromosome 1. Members of this protein family have been implicated in intellectual disability. In summary, this study implicates parent‐of‐origin effects in language impairment, and adds an interesting new dimension to the emerging picture of shared genetic etiology across various neurodevelopmental disorders .     

Application of Human‐Induced Pluripotent Stem Cells (hiPSCs) to Study Synaptopathy of Neurodevelopmental Disorders

Synapses are the basic structural and functional units for information processing and storage in the brain. Their diverse properties and functions ultimately underlie the complexity of human behavior. Proper development and maintenance of synapses are essential for normal functioning of the nervous system. Disruption in synaptogenesis and the consequent alteration in synaptic function have been strongly implicated to cause neurodevelopmental disorders such as autism spectrum disorders (ASDs) and schizophrenia (SCZ). The introduction of human‐induced pluripotent stem cells (hiPSCs) provides a new path to elucidate disease mechanisms and potential therapies. In this review, we will discuss the advantages and limitations of using hiPSC‐derived neurons to study synaptic disorders. Many mutations in genes encoding for proteins that regulate synaptogenesis have been identified in patients with ASDs and SCZ. We use Methyl‐CpG binding protein 2 (MECP2), SH3 and multiple ankyrin repeat domains 3 (SHANK3) and Disrupted in schizophrenia 1 (DISC1) as examples to illustrate the promise of using hiPSCs as cellular models to elucidate the mechanisms underlying disease‐related synaptopathy.     

Development and evaluation of a genome‐wide Coffee 8.5K SNP array and its application for high‐density genetic mapping and for investigating the origin of Coffea arabica L.

Coffee species such as Coffea canephora P. (Robusta) and C. arabica L. (Arabica) are important cash crops in tropical regions around the world. C. arabica is an allotetraploid (2n = 4x = 44) originating from a hybridization event of the two diploid species C. canephora and C. eugenioides (2n = 2x = 22). Interestingly, these progenitor species harbour a greater level of genetic variability and are an important source of genes to broaden the narrow Arabica genetic base. Here, we describe the development, evaluation and use of a single‐nucleotide polymorphism (SNP) array for coffee trees. A total of 8580 unique and informative SNPs were selected from C. canephora and C. arabica sequencing data, with 40% of the SNP located in annotated genes. In particular, this array contains 227 markers associated to 149 genes and traits of agronomic importance. Among these, 7065 SNPs (~82.3%) were scorable and evenly distributed over the genome with a mean distance of 54.4 Kb between markers. With this array, we improved the Robusta high‐density genetic map by adding 1307 SNP markers, whereas 945 SNPs were found segregating in the Arabica mapping progeny. A panel of C. canephora accessions was successfully discriminated and over 70% of the SNP markers were transferable across the three species. Furthermore, the canephora‐derived subgenome of C. arabica was shown to be more closely related to C. canephora accessions from northern Uganda than to other current populations. These validated SNP markers and high‐density genetic maps will be useful to molecular genetics and for innovative approaches in coffee breeding.     

Genotype‐by‐sequencing of the plant‐pathogenic fungi Pyrenophora teres and Sphaerulina musiva utilizing Ion Torrent sequence technology

Genetic and genomics tools to characterize host–pathogen interactions are disproportionately directed to the host because of the focus on resistance. However, understanding the genetics of pathogen virulence is equally important and has been limited by the high cost of de novo genotyping of species with limited marker data. Non‐resource‐prohibitive methods that overcome the limitation of genotyping are now available through genotype‐by‐sequencing (GBS). The use of a two‐enzyme restriction‐associated DNA (RAD)‐GBS method adapted for Ion Torrent sequencing technology provided robust and reproducible high‐density genotyping of several fungal species. A total of 5783 and 2373 unique loci, ‘sequence tags’, containing 16 441 and 9992 single nucleotide polymorphisms (SNPs) were identified and characterized from natural populations of Pyrenophora teres f. maculata and Sphaerulina musiva, respectively. The data generated from the P. teres f. maculata natural population were used in association mapping analysis to map the mating‐type gene to high resolution. To further validate the methodology, a biparental population of P. teres f. teres, previously used to develop a genetic map utilizing simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) markers, was re‐analysed using the SNP markers generated from this protocol. A robust genetic map containing 1393 SNPs on 997 sequence tags spread across 15 linkage groups with anchored reference markers was generated from the P. teres f. teres biparental population. The robust high‐density markers generated using this protocol will allow positional cloning in biparental fungal populations, association mapping of natural fungal populations and population genetics studies.     

Genomic regions affecting backfat thickness and cannon bone circumference identified by genome‐wide association study in a Duroc pig population

We performed a genome‐wide association study using the porcine 60K SNP array to detect QTL regions for nine traits in a three‐generational Duroc samples (n = 651), viz. generations 1, 2 and 3 from a population selected over five generations using a closed nucleus breeding scheme. We applied a linear mixed model for association mapping to detect SNP effects, adjusting for fixed effects (sex and season) and random polygenic effects (reflecting genetic relatedness), and derived a likelihood ratio statistic for each SNP using the efficient mixed‐model association method. We detected a region on SSC6 for backfat thickness (BFT) and on SSC7 for cannon bone circumference (CANNON), with a genome‐wide significance of P < 0.01 after Bonferroni correction. These regions had been detected previously in other pig populations. Six genes are located in the BFT‐associated region, while the CANNON‐associated region includes 66 genes. In the future, significantly associated SNPs, derived by sequencing the coding regions of the six genes in the BFT region, can be used in marker‐assisted selection of BFT, whereas haplotypes constructed from the SSC7 region with strong LD can be used to select for the CANNON trait in our resource family.     

Identifying chromosomal selection‐sweep regions in facial eczema selection‐line animals using an ovine 50K‐SNP array

Facial eczema (FE) is a hepato‐mycotoxicosis found mainly in New Zealand sheep and cattle. When genetics was found to be a factor in FE susceptibility, resistant and susceptible selection lines of Romney sheep were established to enable further investigations of this disease trait. Using the Illumina OvineSNP50 BeadChip, we conducted a selection‐sweep experiment on these FE genetic lines. Two analytical methods were used to detect selection signals, namely the Peddrift test (Dodds & McEwan, 1997) and fixation index F_ST (Weir & Hill, 2002). Of 50 975 single nucleotide polymorphism (SNP) markers tested, there were three that showed highly significant allele frequency differences between the resistant and susceptible animals (Peddrift nominal P < 0.000001). These SNP loci are located on chromosomes OAR1, OAR11 and OAR12 that coincide precisely with the three highest genomic F_ST peaks. In addition, there are nine less significant Peddrift SNPs (nominal P ≤ 0.000009) on OAR6 (n = 2), OAR9 (n = 2), OAR12, OAR19 (n = 2), OAR24 and OAR26. In smoothed F_ST (five‐SNP moving average) plots, the five most prominent peaks are on OAR1, OAR6, OAR7, OAR13 and OAR19. Although these smoothed F_ST peaks do not coincide with the three most significant Peddrift SNP loci, two (on OAR6 and OAR19) overlap with the set of less significant Peddrift SNPs above. Of these 12 Peddrift SNPs and five smoothed F_ST regions, none is close to the FE candidate genes catalase and ABCG2; however, two on OAR1 and one on OAR13 fall within suggestive quantitative trait locus regions identified in a previous genome screen experiment. The present studies indicated that there are at least eight genomic regions that underwent a selection sweep in the FE lines.     

miR‐370 and miR‐373 regulate the pathogenesis of osteoarthritis by modulating one‐carbon metabolism via SHMT‐2 and MECP‐2, respectively

The aim of this study was to determine the mechanism underlying the association between one‐carbon metabolism and DNA methylation during chronic degenerative joint disorder, osteoarthritis (OA). Articular chondrocytes were isolated from human OA cartilage and normal cartilage biopsied, and the degree of cartilage degradation was determined by safranin O staining. We found that the expression levels of SHMT‐2 and MECP‐2 were increased in OA chondrocytes, and 3′UTR reporter assays showed that SHMT‐2 and MECP‐2 are the direct targets of miR‐370 and miR‐373, respectively, in human articular chondrocytes. Our experiments showed that miR‐370 and miR‐373 levels were significantly lower in OA chondrocytes compared to normal chondrocytes. Overexpression of miR‐370 or miR‐373, or knockdown of SHMT‐2 or MECP‐2 reduced both MMP‐13 expression and apoptotic cell death in cultured OA chondrocytes. In vivo, we found that introduction of miR‐370 or miR‐373 into the cartilage of mice that had undergone destabilization of the medial meniscus (DMM) surgery significantly reduced the cartilage destruction in this model, whereas introduction of SHMT‐2 or MECP‐2 increased the severity of cartilage destruction. Together, these results show that miR‐370 and miR‐373 contribute to the pathogenesis of OA and act as negative regulators of SHMT‐2 and MECP‐2, respectively.     

A genome‐wide association study for the incidence of persistent bovine viral diarrhea virus infection in cattle

Bovine viral diarrhea viruses (BVDV) comprise a diverse group of viruses that cause disease in cattle. BVDV may establish both transient and persistent infections depending on the developmental stage of the animal at exposure. The objective was to determine whether genomic regions harboring single nucleotide polymorphisms (SNPs) could be associated with the presence or absence of persistent BVDV infection. A genome‐wide association approach based on 777 000 SNP markers was used. Samples of animals identified as positive (n = 1200) or negative (n = 1200) for the presence of BVDV in skin samples (n = 1200) were used. DNA samples were combined in 24 pools (100 animals per pool). One SNP, significant at the 5 percent genome‐wide level (P = 9.41 × 10^?8), was detected on chromosome 14, located at position 80 675 176 bp. Fifteen SNPs, residing on chromosomes 1, 2, 6, 8, 10, 15 and 18, were moderately associated (P < 1 × 10^?5) with persistent BVDV infection. Results show that genes harboring or neighboring significant SNPs are involved in leucopenia, signal transduction, RNA splicing and DNA methylation processes.     

Genome‐wide association study of body morphological traits in Sudanese goats

Long‐term selection of goats for a certain production system and/or different environmental conditions will be reflected in the body morphology of the animals under selection. To investigate the variation contributing to different morphological traits and to identify genomic regions that are associated with body morphological traits in Sudanese goats, we genotyped 96 females belonging to four Sudanese goat breeds with the SNP52 BeadChip. After quality control of the data, the genome‐wide association study was performed using 95 goats and 24 027 informative single nucleotide polymorphisms (SNPs). Bicoastal diameter was significantly associated (LOD = 6.32) with snp10185‐scaffold1365‐620922 on chromosome 2. The minor allele has an additive effect, increasing the bicoastal diameter by 2.6 cm. A second significant association was found between body length and snp56482‐scaffold89‐467312 on chromosome 3 (LOD = 5.65). The minor allele is associated with increased body length. Additionally, five regions were suggestive for cannon bone, head width, rump length and withers height (LOD > 5). Only one gene (CNTNAP5) is located within the 1‐Mb region surrounding the significant SNP for bicoastal diameter on chromosome 2. The body length QTL on chromosome 3 harbors 49 genes. Further research is required to validate the observed associations and to prioritize candidate genes.     

Novel SNPs of the bovine <Emphasis Type="Italic">CACNA2D1</Emphasis> gene and their association with carcass and meat quality traits

Calcium channel, voltage-dependent, alpha-2/delta subunit 1 (CACNA2D1) gene encodes a member of the alpha-2/delta subunit family, proteins are accessory molecules associated with voltage-gated calcium channels, and increase the density at the plasma membrane of calcium channels activated by high voltage. The main objective of the present study was to identify polymorphisms of CACNA2D1 gene, and to analyze associations between these polymorphisms and carcass and meat quality traits in cattle. In this study, through PCR-SSCP and DNA sequencing methods, two new allelic variant corresponding to the C → G and G → T mutations at positions 526740 and 537917 in the exon25 and exon35 of bovine CACNA2D1 gene, respectively, could be detected. SNP C526740G is a nonsynonymous mutation, resulting in a Cysteine (Cys) to Tryptophan (Trp) amino acid replacement and SNP G537917T resulting in an Aspartic (Asp) to Tyrosine (Tyr) amino acid replacement. The gene-specific SNP markers association analysis was investigated. The C526740G was significantly associated with Meat color (MC) (P = 0.0297) and Backfat thickness (BF) (P < 0.001). The G537917A indicated significant association with Dressing percentage (DP) (P = 0.0485). No significant association, however, was detected between any of the marker genotype and other traits measured in this study. Results from this study initially suggested that CACNA2D1 gene is one of the potential candidate genes influencing carcass and meat quality traits and gene-specific SNPs may be a useful marker for MAS programs in cattle breeding.