LD-CNV: rapid and simple discovery of chromosomal translocations using linkage disequilibrium between copy number variable loci.

Large-scale structural variations, such as chromosomal translocations, can have profound effects on fitness and phenotype, but are difficult to identify and characterize. Here, we describe a simple and effective method aimed at identifying translocations using only the dosage of sequence reads mapped on the reference genome. We binned reads on genomic segments sized according to sequencing coverage and identified instances when copy number segregated in populations. For each dosage-polymorphic 1 Mb bin, we tested independence, effectively an apparent linkage disequilibrium (LD), with other variable bins. In nine potato (Solanum tuberosum) dihaploid families translocations affecting pericentromeric regions were common and in two cases were due to genomic misassembly. In two populations, we found evidence for translocation affecting euchromatic arms. In cv. PI 310467, a nonreciprocal translocation between chromosomes (chr.) 7 and 8 resulted in a 5-3 copy number change affecting several Mb at the respective chromosome tips. In cv. "Alca Tarma," the terminal arm of chr. 4 translocated to the tip of chr. 1. Using oligonucleotide-based fluorescent in situ hybridization painting probes (oligo-FISH), we tested and confirmed the predicted arrangement in PI 310467. In 192 natural accessions of Arabidopsis thaliana, dosage haplotypes tended to vary continuously and resulted in higher noise, while apparent LD between pericentromeric regions suggested the effect of repeats. This method, LD-CNV, should be useful in species where translocations are suspected because it tests linkage without the need for genotyping.

Genetic diversity and population structure of Alternaria species from tomato and potato in North Carolina and Wisconsin.

Early blight (EB) caused by Alternaria linariae or Alternaria solani and leaf blight (LB) caused by A. alternata are economically important diseases of tomato and potato. Little is known about the genetic diversity and population structure of these pathogens in the United States. A total of 214 isolates of A. alternata (n = 61), A. linariae (n = 96), and A. solani (n = 57) were collected from tomato and potato in North Carolina and Wisconsin and grouped into populations based on geographic locations and tomato varieties. We exploited 220 single nucleotide polymorphisms derived from DNA sequences of 10 microsatellite loci to analyse the population genetic structure between species and between populations within species and infer the mode of reproduction. High genetic variation and genotypic diversity were observed in all the populations analysed. The null hypothesis of the clonality test based on the index of association [Formula: see text] was rejected, and equal frequencies of mating types under random mating were detected in some studied populations of Alternaria spp., suggesting that recombination can play an important role in the evolution of these pathogens. Most genetic differences were found between species, and the results showed three distinct genetic clusters corresponding to the three Alternaria spp. We found no evidence for clustering of geographic location populations or tomato variety populations. Analyses of molecular variance revealed high (> 85%) genetic variation within individuals in a population, confirming a lack of population subdivision within species. Alternaria linariae populations harboured more multilocus genotypes (MLGs) than A. alternata and A. solani populations and shared the same MLG between populations within a species, which was suggestive of gene flow and population expansion. Although both A. linariae and A. solani can cause EB on tomatoes and potatoes, these two species are genetically differentiated. Our results provide new insights into the evolution and structure of Alternaria spp. and can lead to new directions in optimizing management strategies to mitigate the impact of these pathogens on tomato and potato production in North Carolina and Wisconsin.

The genetic architecture of the human thalamus and its overlap with ten common brain disorders.

The thalamus is a vital communication hub in the center of the brain and consists of distinct nuclei critical for consciousness and higher-order cortical functions. Structural and functional thalamic alterations are involved in the pathogenesis of common brain disorders, yet the genetic architecture of the thalamus remains largely unknown. Here, using brain scans and genotype data from 30,114 individuals, we identify 55 lead single nucleotide polymorphisms (SNPs) within 42 genetic loci and 391 genes associated with volumes of the thalamus and its nuclei. In an independent validation sample (n = 5173) 53 out of the 55 lead SNPs of the discovery sample show the same effect direction (sign test, P = 8.6e-14). We map the genetic relationship between thalamic nuclei and 180 cerebral cortical areas and find overlapping genetic architectures consistent with thalamocortical connectivity. Pleiotropy analyses between thalamic volumes and ten psychiatric and neurological disorders reveal shared variants for all disorders. Together, these analyses identify genetic loci linked to thalamic nuclei and substantiate the emerging view of the thalamus having central roles in cortical functioning and common brain disorders.

Population structure and genome-wide association studies in bread wheat for phosphorus efficiency traits using 35 K Wheat Breeder's Affymetrix array.

Soil bioavailability of phosphorus (P) is a major concern for crop productivity worldwide. As phosphatic fertilizers are a non-renewable resource associated with economic and environmental issues so, the sustainable option is to develop P use efficient crop varieties. We phenotyped 82 diverse wheat (Triticum aestivum L.) accessions in soil and hydroponics at low and sufficient P. To identify the genic regions for P efficiency traits, the accessions were genotyped using the 35 K-SNP array and genome-wide association study (GWAS) was performed. The high-quality SNPs across the genomes were evenly distributed with polymorphic information content values varying between 0.090 and 0.375. Structure analysis revealed three subpopulations (C1, C2, C3) and the phenotypic responses of these subpopulations were assessed for P efficiency traits. The C2 subpopulation showed the highest genetic variance and heritability values for numerous agronomically important traits as well as strong correlation under both P levels in soil and hydroponics. GWAS revealed 78 marker-trait associations (MTAs) but only 35 MTAs passed Bonferroni Correction. A total of 297 candidate genes were identified for these MTAs and their annotation suggested their involvement in several biological process. Out of 35, nine (9) MTAs were controlling polygenic trait (two controlling four traits, one controlling three traits and six controlling two traits). These multi-trait MTAs (each controlling two or more than two correlated traits) could be utilized for improving bread wheat to tolerate low P stress through marker-assisted selection (MAS).

Genetic analysis of a potato (Solanum tuberosum L.) breeding collection for southern Colombia using Single Nucleotide Polymorphism (SNP) markers.

Detailed knowledge on genetic parameters such as diversity, structure, and linkage disequilibrium (LD) and identification of duplicates in a germplasm bank and/or breeding collection are essential to conservation and breeding strategies in any crop. Therefore, the potato genetic breeding collection at the Universidad de Nariño in Colombia, which is made up of diploid and tetraploid genotypes in two of the more diverse genebanks in the world, was analyzed with 8303 single nucleotide polymorphisms (SNP) from SolCAP version 1. In total, 144 genotypes from this collection were analyzed identifying an 57.2% of the polymorphic markers that allowed establishing two and three subpopulations that differentiated the diploid genotypes from the tetraploids. These subpopulations had high levels of heterozygosity and linkage disequilibrium. The diversity levels were higher in the tetraploid genotypes, while the LD levels were higher in the diploid genotypes. For the tetraploids, the genotypes from Peru had greater diversity and lower linkage disequilibrium than those from Colombia, which had slightly lower diversity and higher degrees of LD. The genetic analysis identified, adjusted and/or selected diploid and tetraploid genotypes under the following characteristics: 1) errors in classification associated with the level of ploidy; 2) presence of duplicates; and 3) genotypes with broad genetic distances and potential use in controlled hybridization processes. These analyses suggested that the potato genetic breeding collection at the Universidad de Nariño has a genetic base with a potential use in breeding programs for this crop in the Department of Nariño, in southern Colombia.

Association mapping and genomic selection for sorghum adaptation to tropical soils of Brazil in a sorghum multiparental random mating population.

KEY MESSAGE: A multiparental random mating population used in sorghum breeding is amenable for the detection of QTLs related to tropical soil adaptation, fine mapping of underlying genes and genomic selection approaches. Tropical soils where low phosphorus (P) and aluminum (Al) toxicity limit sorghum [Sorghum bicolor (L.) Moench] production are widespread in the developing world. We report on BRP13R, a multiparental random mating population (MP-RMP), which is commonly used in sorghum recurrent selection targeting tropical soil adaptation. Recombination dissipated much of BRP13R's likely original population structure and average linkage disequilibrium (LD) persisted up to 2.5 Mb, establishing BRP13R as a middle ground between biparental populations and sorghum association panels. Genome-wide association mapping (GWAS) identified conserved QTL from previous studies, such as for root morphology and grain yield under low-P, and indicated the importance of dominance in the genetic architecture of grain yield. By overlapping consensus QTL regions, we mapped two candidate P efficiency genes to a ~ 5 Mb region on chromosomes 6 (ALMT) and 9 (PHO2). Remarkably, we find that only 200 progeny genotyped with ~ 45,000 markers in BRP13R can lead to GWAS-based positional cloning of naturally rare, subpopulation-specific alleles, such as for SbMATE-conditioned Al tolerance. Genomic selection was found to be useful in such MP-RMP, particularly if markers in LD with major genes are fitted as fixed effects into GBLUP models accommodating dominance. Shifts in allele frequencies in progeny contrasting for grain yield indicated that intermediate to minor-effect genes on P efficiency, such as SbPSTOL1 genes, can be employed in pre-breeding via allele mining in the base population. Therefore, MP-RMPs such as BRP13R emerge as multipurpose resources for efficient gene discovery and deployment for breeding sorghum cultivars adapted to tropical soils.

Higher 25-hydroxyvitamin D level is associated with increased risk for Behçet's disease.

BACKGROUND & AIM: Previous studies showed a vitamin D deficiency in patients with Behçet's disease, suggesting potential benefits of vitamin D supplementation in the prevention and treatment of Behçet's disease. Interpretation of these studies may be limited by reverse causality or confounding bias. We aim to determine the causal association between serum 25-hydroxyvitamin D [25(OH)D] and the risk of Behçet's disease by Mendelian randomization. METHODS: An allele score formed by four variants (rs2282679, rs10741657, rs12785878 and rs6013897) that were associated with serum 25(OH)D level, was examined using data of genome-wide association study (GWAS) on 999 Behçet's disease and 4417 healthy individuals of Chinese ancestry and validated using data of GWAS on 1215 Behçet's disease and 1278 controls of Turkish ancestry. The primary outcome was the risk of Behçet's disease, evaluated by an inverse variance weighted average of the associations with genetically determined 25(OH)D levels. RESULTS: The inverse variance weighted estimate showed that genetically increased 25(OH)D level was associated with a higher risk of Behçet's disease. In the Chinese cohort, the odds ratio for Behçet's disease in one standard deviation increase of natural log-transformed 25(OH)D level was 3.82 (95% CI: 1.27-11.42). Data from Turkish cohort confirmed the association with Behçet's disease (OR, 95% CI: 4.18, 1.15-15.12). In overall combination of Chinese and Turkish cohorts, the odds ratio for Behçet's disease per standard deviation increase of natural log-transformed 25(OH)D level was estimated to be 3.96 (95% CI: 1.72-9.13; P = 0.001). No significant evidence of pleiotropy and heterogeneity was detected. CONCLUSIONS: On the basis of evidence in 7909 human beings, this study provides the newest indication that a lifelong higher 25(OH)D level is associated with an increased risk of Behçet's disease. Special attention should be paid to the potential harm of long-term or high-dose use of vitamin D supplements in clinical practice.

Variant-to-Gene-Mapping Analyses Reveal a Role for the Hypothalamus in Genetic Susceptibility to Inflammatory Bowel Disease.

BACKGROUND & AIMS: Inflammatory bowel disease (IBD) is a polygenic disorder characterized principally by dysregulated inflammation impacting the gastrointestinal tract. However, there also is increasing evidence for a clinical association with stress and depression. Given the role of the hypothalamus in stress responses and in the pathogenesis of depression, useful insights could be gleaned from understanding its genetic role in IBD. METHODS: We conducted genetic correlation analyses on publicly available genome-wide association study summary statistics for depression and IBD traits to identify genetic commonalities. We used partitioned linkage disequilibrium score regression, leveraging our ATAC sequencing and promoter-focused Capture C data, to measure enrichment of IBD single-nucleotide polymorphisms within promoter-interacting open chromatin regions of human embryonic stem cell-derived hypothalamic-like neurons (HNs). Using the same data sets, we performed variant-to-gene mapping to implicate putative IBD effector genes in HNs. To contrast these results, we similarly analyzed 3-dimensional genomic data generated in epithelium-derived colonoids from rectal biopsy specimens from donors without pathologic disease noted at the time of colonoscopy. Finally, we conducted enrichment pathway analyses on the implicated genes to identify putative IBD dysfunctional pathways. RESULTS: We found significant genetic correlations (rg) of 0.122 with an adjusted P (Padj) = 1.4 × 10-4 for IBD: rg = 0.122; Padj = 2.5 × 10-3 for ulcerative colitis and genetic correlation (rg) = 0.094; Padj = 2.5 × 10-3 for Crohn's disease, and significant approximately 4-fold (P = .005) and approximately 7-fold (P = .03) enrichment of IBD single-nucleotide polymorphisms in HNs and colonoids, respectively. We implicated 25 associated genes in HNs, among which CREM, CNTF, and RHOA encode key regulators of stress. Seven genes also additionally were implicated in the colonoids. We observed an overall enrichment for immune and hormonal signaling pathways, and a colonoid-specific enrichment for microbiota-relevant terms. CONCLUSIONS: Our results suggest that the hypothalamus warrants further study in the context of IBD pathogenesis.

Identification of genetic factors controlling phosphorus utilization efficiency in wheat by genome-wide association study with principal component analysis.

Despite the economic importance of P utilization efficiency, information on genetic factors underlying this trait remains elusive. To address that, we performed a genome-wide association study in a spring wheat diversity panel ranging from landraces to elite varieties. We evaluated the phenotype variation for P utilization efficiency in controlled conditions and genotype variation using wheat 90 K SNP array. Phenotype variables were transformed into a smaller set of uncorrelated principal components that captured the most important variation data. We identified two significant loci associated with both P utilization efficiency and the 1st principal component on chromosomes 3A and 4A: qPE1-3A and qPE2-4A. Annotation of genes at these loci revealed 53 wheat genes, among which 6 were identified in significantly enriched pathways. The expression pattern of these 6 genes indicated that TraesCS4A02G481800, involved in pyruvate metabolism and TCA cycle, had a significantly higher expression in the P efficient variety under limited P conditions. Further characterization of these loci and candidate genes can help stimulate P utilization efficiency in wheat.

Genomic predictions and genome-wide association studies based on RAD-seq of quality-related metabolites for the genomics-assisted breeding of tea plants.

Effectively using genomic information greatly accelerates conventional breeding and applying it to long-lived crops promotes the conversion to genomic breeding. Because tea plants are bred using conventional methods, we evaluated the potential of genomic predictions (GPs) and genome-wide association studies (GWASs) for the genetic breeding of tea quality-related metabolites using genome-wide single nucleotide polymorphisms (SNPs) detected from restriction site-associated DNA sequencing of 150 tea accessions. The present GP, based on genome-wide SNPs, and six models produced moderate prediction accuracy values (r) for the levels of most catechins, represented by ( -)-epigallocatechin gallate (r = 0.32-0.41) and caffeine (r = 0.44-0.51), but low r values for free amino acids and chlorophylls. Integrated analysis of GWAS and GP detected potential candidate genes for each metabolite using 80-160 top-ranked SNPs that resulted in the maximum cumulative prediction value. Applying GPs and GWASs to tea accession traits will contribute to genomics-assisted tea breeding.

Phenotypic and genetic variation in phosphorus-deficiency-tolerance traits in Chinese wheat landraces.

BACKGROUND: Phosphorus deficiency is a major limiting factors for affecting crop production globally. To understand the genetic variation of phosphorus-deficiency-tolerance, a total of 15 seedling traits were evaluated among 707 Chinese wheat landraces under application of phosphorus (AP) and non-application of phosphorus (NP). A total of 18,594 single-nucleotide polymorphisms and 38,678 diversity arrays technology sequencing markers were used to detect marker-trait associations under AP and NP. RESULTS: Top ten genotypes with extremely tolerance and bottommost ten genotypes with extremely sensitivity were selected from 707 Chinese wheat landraces for future breeding and genetic analysis. A total of 55 significant markers (81 marker-trait associations) for 13 traits by both CMLM and SUPER method. These were distributed on chromosomes 1A, 1B, 2A, 2B, 2D, 3A, 4B, 5A, 5B, 6A, 6B, 6D, 7A and 7B. Considering the linkage disequilibrium decay distance, 25 and 12 quantitative trait loci (QTL) were detected under AP and NP, respectively (9 QTL were specific to NP). CONCLUSIONS: The extremely tolerant landraces could be used for breeding phosphorus-deficiency-tolerant cultivars. The QTL could be useful in wheat breeding through marker-assisted selection. Our findings provide new insight into the genetic analysis of P-deficiency-tolerance, and will be helpful for breeding P-deficiency-tolerant cultivars.

Representation of CYP3A4, CYP3A5 and UGT1A4 Polymorphisms within Croatian Breast Cancer Patients' Population.

Single nucleotide polymorphism (SNP) in genes encoding drug-metabolizing enzymes (DME) could have a critical role in individual responses to anastrozole. Frequency of CYP3A4*1B, CYP3A5*3 and UGT1A4*2 SNPs in 126 Croatian breast cancer (BC) patients and possible association with anastrozole-induced undesirable side effects were analyzed. Eighty-two postmenopausal patients with estrogen receptor (ER)-positive BC treated with anastrozole and 44 postmenopausal ER-positive BC patients before hormonal adjuvant therapy were included in the study. Genomic DNA was genotyped by TaqMan Real-Time PCR. Bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry. The homozygotes for the variant G allele of CYP3A5*3 were predominant (88%), and the homozygotes for the reference A allele were not detected. While homozygotes for the variant G allele of CYP3A4*1B were not detected, predominantly wild type homozygotes for A allele (94%) were present. CYP3A4*1B and CYP3A5*3 SNPs were in 84.3% linkage disequilibrium (D' = 0.843) and 95.1% (D' = 0.951) in group treated with anastrozole and w/o treatment, respectively. Homozygotes for the A allele of UGT1A4*2 were not detected in our study groups. Although the variant CYP3A5*3 allele, which might result in poor metabolizer phenotype and more pronounced side effects, was predominant, significant association with BMD changes induced by anastrozole were not confirmed.

Genetic diversity among cultivated beets (Beta vulgaris) assessed via population-based whole genome sequences.

BACKGROUND: Diversification on the basis of utilization is a hallmark of Beta vulgaris (beet), as well as other crop species. Often, crop improvement and management activities are segregated by crop type, thus preserving unique genome diversity and organization. Full interfertility is typically retained in crosses between these groups and more traits may be accessible if the genetic basis of crop type lineage were known, along with available genetic markers to effect efficient transfer (e.g., via backcrossing). Beta vulgaris L. (2n =18) is a species complex composed of diverged lineages (e.g., crop types), including the familiar table, leaf (chard), fodder, and sugar beet crop types. Using population genetic and statistical methods with whole genome sequence data from pooled samples of 23 beet cultivars and breeding lines, relationships were determined between accessions based on identity-by-state metrics and shared genetic variation among lineages. RESULTS: Distribution of genetic variation within and between crop types showed extensive shared (e.g. non-unique) genetic variation. Lineage specific variation (e.g. apomorphy) within crop types supported a shared demographic history within each crop type, while principal components analysis revealed strong crop type differentiation. Relative contributions of specific chromosomes to genome wide differentiation were ascertained, with each chromosome revealing a different pattern of differentiation with respect to crop type. Inferred population size history for each crop type helped integrate selection history for each lineage, and highlighted potential genetic bottlenecks in the development of cultivated beet lineages. CONCLUSIONS: A complex evolutionary history of cultigroups in Beta vulgaris was demonstrated, involving lineage divergence as a result of selection and reproductive isolation. Clear delineation of crop types was obfuscated by historical gene flow and common ancestry (e.g. admixture and introgression, and sorting of ancestral polymorphism) which served to share genome variation between crop types and, likely, important phenotypic characters. Table beet was well differentiated as a crop type, and shared more genetic variation within than among crop types. The sugar beet group was not quite as well differentiated as the table beet group. Fodder and chard groups were intermediate between table and sugar groups, perhaps the result of less intensive selection for end use.

Distribution of P1(D1) wart disease resistance in potato germplasm and GWAS identification of haplotype-specific SNP markers.

KEY MESSAGE: A Genome-Wide Association Study using 330 commercial potato varieties identified haplotype specific SNP markers associated with pathotype 1(D1) wart disease resistance. Synchytrium endobioticum is a soilborne obligate biotrophic fungus responsible for wart disease. Growing resistant varieties is the most effective way to manage the disease. This paper addresses the challenge to apply molecular markers in potato breeding. Although markers linked to Sen1 were published before, the identification of haplotype-specific single-nucleotide polymorphisms may result in marker assays with high diagnostic value. To identify hs-SNP markers, we performed a genome-wide association study (GWAS) in a panel of 330 potato varieties representative of the commercial potato gene pool. SNP markers significantly associated with pathotype 1 resistance were identified on chromosome 11, at the position of the previously identified Sen1 locus. Haplotype specificity of the SNP markers was examined through the analysis of false positives and false negatives and validated in two independent full-sib populations. This paper illustrates why it is not always feasible to design markers without false positives and false negatives for marker-assisted selection. In the case of Sen1, founders could not be traced because of a lack of identity by descent and because of the decay of linkage disequilibrium between Sen1 and flanking SNP markers. Sen1 appeared to be the main source of pathotype 1 resistance in potato varieties, but it does not explain all the resistance observed. Recombination and introgression breeding may have introduced new, albeit rare haplotypes involved in pathotype 1 resistance. The GWAS approach, in such case, is instrumental to identify SNPs with the best possible diagnostic value for marker-assisted breeding.

Natural variation and selection in GmSWEET39 affect soybean seed oil content.

Soybean (Glycine max) is a major contributor to the world oilseed production. Its seed oil content has been increased through soybean domestication and improvement. However, the genes underlying the selection are largely unknown. The present contribution analyzed the expression patterns of genes in the seed oil quantitative trait loci with strong selective sweep signals, then used association, functional study and population genetics to reveal a sucrose efflux transporter gene, GmSWEET39, controlling soybean seed oil content and under selection. GmSWEET39 is highly expressed in soybean seeds and encodes a plasma membrane-localized protein. Its expression level is positively correlated with soybean seed oil content. The variation in its promoter and coding sequence leads to different natural alleles of this gene. The GmSWEET39 allelic effects on total oil content were confirmed in the seeds of soybean recombinant inbred lines, transgenic Arabidopsis, and transgenic soybean hairy roots. The frequencies of its superior alleles increased from wild soybean to cultivated soybean, and are much higher in released soybean cultivars. The findings herein suggest that the sequence variation in GmSWEET39 affects its relative expression and oil content in soybean seeds, and GmSWEET39 has been selected to increase seed oil content during soybean domestication and improvement.

Genome wide association study (GWAS) and identification of candidate genes for yield and oil yield related traits in oil palm (Eleaeis guineensis) using SNPs by genotyping-based sequencing.

The genotyping-based sequencing (GBS) method used for GWAS of four yield and seven oil yield related traits on highly diverse African oil palm germplasm. GBS generated 325 million-reads covering 50.78Gb of sequence data, with an average of 3.4 million-reads per sample. Finally, 4031 fully informative SNPs with a range between 157 on chromosome 15 to 455 on chromosome 1 were used for GWAS. Association mapping resulted in identification of 40 highly significant loci, where more genetic loci were found to be associated with oil to bunch (OB), followed by average bunch weight (ABW). The loci, SGI|593,593|linked to QTNOB3 explained high amount of phenotypic variance (25.3%). The nucleotide sequences of linked genetic loci for OB were found to be similar to mitogen activated protein kinase-5 (MAPK-5) protein which is an early flowering protein. The significant loci identified can be used to select desirable palms at early stage through marker assisted selection.

A Combined Linkage and GWAS Analysis Identifies QTLs Linked to Soybean Seed Protein and Oil Content.

Soybean is an excellent source of vegetable protein and edible oil. Understanding the genetic basis of protein and oil content will improve the breeding programs for soybean. Linkage analysis and genome-wide association study (GWAS) tools were combined to detect quantitative trait loci (QTL) that are associated with protein and oil content in soybean. Three hundred and eight recombinant inbred lines (RILs) containing 3454 single nucleotide polymorphism (SNP) markers and 200 soybean accessions, including 94,462 SNPs and indels, were applied to identify QTL intervals and significant SNP loci. Intervals on chromosomes 1, 15, and 20 were correlated with both traits, and QTL qPro15-1, qPro20-1, and qOil5-1 reproducibly correlated with large phenotypic variations. SNP loci on chromosome 20 that overlapped with qPro20-1 were reproducibly connected to both traits by GWAS (p < 10-4). Twenty-five candidate genes with putative roles in protein and/or oil metabolisms within two regions (qPro15-1, qPro20-1) were identified, and eight of these genes showed differential expressions in parent lines during late reproductive growth stages, consistent with a role in controlling protein and oil content. The new well-defined QTL should significantly improve molecular breeding programs, and the identified candidate genes may help elucidate the mechanisms of protein and oil biosynthesis.

Disruption of an enhancer associated with addictive behaviour within the cannabinoid receptor-1 gene suggests a possible role in alcohol intake, cannabinoid response and anxiety-related behaviour.

The cannabinoid-1 receptor (CB1) plays a critical role in a number of biological processes including nutrient intake, addiction and anxiety-related behaviour. Numerous studies have shown that expression of the gene encoding CB1 (CNR1) is highly dynamic with changes in the tissue specific expression of CNR1 associated with brain homeostasis and disease progression. However, little is known of the mechanisms regulating this dynamic expression. To gain a better understanding of the genomic mechanisms modulating the expression of CNR1 in health and disease we characterised the role of a highly conserved regulatory sequence (ECR1) in CNR1 intron 2 that contained a polymorphism in linkage disequilibrium with disease associated SNPs. We used CRISPR/CAS9 technology to disrupt ECR1 within the mouse genome. Disruption of ECR1 significantly reduced CNR1 expression in the hippocampus but not in the hypothalamus. These mice also displayed an altered sex-specific anxiety-related behavioural profile (open field test), reduced ethanol intake and a reduced hypothermic response following CB1 agonism. However, no significant changes in feeding patterns were detected. These data suggest that, whilst not all of the expression of CNR1 is modulated by ECR1, this highly conserved enhancer is required for appropriate physiological responses to a number of stimuli. The combination of comparative genomics and CRISPR/CAS9 disruption used in our study to determine the functional effects of genetic and epigenetic changes on the activity of tissue-specific regulatory elements at the CNR1 locus represent an important first step in gaining a mechanistic understanding of cannabinoid regulatory pharmacogenetics.

Genetic diversity, linkage disequilibrium, and population structure analysis of the tea plant (Camellia sinensis) from an origin center, Guizhou plateau, using genome-wide SNPs developed by genotyping-by-sequencing.

BACKGROUND: To efficiently protect and exploit germplasm resources for marker development and breeding purposes, we must accurately depict the features of the tea populations. This study focuses on the Camellia sinensis (C. sinensis) population and aims to (i) identify single nucleotide polymorphisms (SNPs) on the genome level, (ii) investigate the genetic diversity and population structure, and (iii) characterize the linkage disequilibrium (LD) pattern to facilitate next genome-wide association mapping and marker-assisted selection. RESULTS: We collected 415 tea accessions from the Origin Center and analyzed the genetic diversity, population structure and LD pattern using the genotyping-by-sequencing (GBS) approach. A total of 79,016 high-quality SNPs were identified; the polymorphism information content (PIC) and genetic diversity (GD) based on these SNPs showed a higher level of genetic diversity in cultivated type than in wild type. The 415 accessions were clustered into three groups by STRUCTURE software and confirmed using principal component analyses (PCA)-wild type, cultivated type, and admixed wild type. However, unweighted pair group method with arithmetic mean (UPGMA) trees indicated the accessions should be grouped into more clusters. Further analyses identified four groups, the Pure Wild Type, Admixed Wild Type, ancient landraces and modern landraces using STRUCTURE, and the results were confirmed by PCA and UPGMA tree method. A higher level of genetic diversity was detected in ancient landraces and Admixed Wild Type than that in the Pure Wild Type and modern landraces. The highest differentiation was between the Pure Wild Type and modern landraces. A relatively fast LD decay with a short range (kb) was observed, and the LD decays of four inferred populations were different. CONCLUSIONS: This study is, to our knowledge, the first population genetic analysis of tea germplasm from the Origin Center, Guizhou Plateau, using GBS. The LD pattern, population structure and genetic differentiation of the tea population revealed by our study will benefit further genetic studies, germplasm protection, and breeding.

Assessment of Genetic Differentiation and Linkage Disequilibrium in Solanum pimpinellifolium Using Genome-Wide High-Density SNP Markers.

To mine new favorable alleles for tomato breeding, we investigated the feasibility of utilizing Solanum pimpinellifolium as a diverse panel of genome-wide association study through the restriction site-associated DNA sequencing technique. Previous attempts to conduct genome-wide association studies using S. pimpinellifolium were impeded by an inability to correct for population stratification and by lack of high-density markers to address the issue of rapid linkage disequilibrium decay. In the current study, a set of 24,330 SNPs was identified using 99 S. pimpinellifolium accessions from the Tomato Genetic Resource Center. Approximately 84% of PstI site-associated DNA sequencing regions were located in the euchromatic regions, resulting in the tagging of most SNPs on or near genes. Our genotypic data suggested that S. pimpinellifolium were divided into three single-ancestry subpopulations and four mixed-ancestry subpopulations. Additionally, our SNP genotypic data consistently confirmed the genetic differentiation, achieving a relatively reliable correction of population stratification. Previous studies utilized the 8K tomato SNP array, SolCAP, to investigate the genetic variation of S. pimpinellifolium and we performed a meta-analysis of these genotypes. The result suggested SolCAP array was less appropriate to profile the genetic differentiation of S. pimpinellifolium when more accessions were involved because the samples belonging to the same accession demonstrated different genome patterns. Moreover, as expected, rapid linkage disequilibrium decay was observed in S. pimpinellifolium, especially in euchromatic regions. Approximately two-thirds of the flanking SNP markers did not display linkage disequilibrium based on r2 = 0.1. However, the 18-Kb linkage disequilibrium decay indeed reveals the potential of single-gene resolution in GWAS when markers are saturated.