Unique Loci Research Articles

Parental Reconstruction from a Half-Sib Population of Stoneless Jujube Ziziphus jujuba Mill. Based on Individual Specific SNP Markers Using Multiplex PCR.

The selection of unique and individual-specific SNPs is important as compared with universal SNPs for individual identification. Therefore, the main significance of this research is the selection of specific SNPs in male parent and the identification of offspring with these specific SNPs in their genome by multiplex PCR, which is utilized for genotyping of 332 half-sib plants of Ziziphus jujuba.This cost-effective method makes as much as possible to utilize the same amount of each pair of various targeted loci primers. After PCR amplification of targeted genome parts, the mixed products can be directly used in a next-generation sequencing platform. We concomitantly amplified 10 unique SNP loci at 10 different chromosomes of male JingZao 39 plants in 332 half-sib plants and sequenced them on the Illumina Novaseq 6000 platform. Analysis of SNP genotyping accuracy of 332 half-sib plants showed that all 10 unique SNPs in all 332 plants were correctly amplified in this multiplex PCR method. Furthermore, based on Mendelian inheritance, we identified 124 full-sib plants that have 10 unique SNPs in their genomes. These results were further confirmed by whole genome resequencing of 82 randomly selected half-sib plants, and the identity-by-descent values of all full-sib plants were between 0.4399 to 0.5652. This study displayed a cost-effective multiplex PCR method and proper identification of pollen parent through specific SNPs in half-sib progenies and firstly obtained a full-sib population between 'Wuhezao' and 'JingZao 39', segregating for stone and stoneless fruit.

Genome-Wide Association Study of Sweet Potato Storage Root Traits Using GWASpoly, a Gene Dosage-Sensitive Model.

Sweet potato (Ipomoea batatas) is an important food crop that plays a pivotal role in preserving worldwide food security. Due to its polyploid genome, high heterogeneity, and phenotypic plasticity, sweet potato genetic characterization and breeding is challenging. Genome-wide association studies (GWASs) can provide important resources for breeders to improve breeding efficiency and effectiveness. GWASpoly was used to identify 28 single nucleotide polymorphisms (SNPs), comprising 21 unique genetic loci, associated with sweet potato storage root traits including dry matter (4 loci), subjective flesh color (5 loci), flesh hue angle (3 loci), and subjective skin color and skin hue angle (9 loci), in 384 accessions from the USDA sweet potato germplasm collection. The I. batatas 'Beauregard' and I. trifida reference genomes were utilized to identify candidate genes located within 100 kb from the SNPs that may affect the storage traits of dry matter, flesh color, and skin color. These candidate genes include transcription factors (especially Myb, bHLH, and WRKY family members), metabolite transporters, and metabolic enzymes and associated proteins involved in starch, carotenoid, and anthocyanin synthesis. A greater understanding of the genetic loci underlying sweet potato storage root traits will enable marker-assisted breeding of new varieties with desired traits. This study not only reinforces previous research findings on genes associated with dry matter and β-carotene content but also introduces novel genetic loci linked to these traits as well as other root characteristics.

Development of a haplotype reference panel and analysis of genetic structure in Golden Pompano (Trachinotus ovatus)

Trachinotus ovatus, a valuable marine fish, is primarily farmed in the South China Sea. Intensive aquaculture can lead to genetic uniformity and disease risks. Additionally, the high cost of genome sequencing restricts knowledge of genetic variation, hindering genetic analysis and breeding advancements. This study investigated the genetic diversity and structure of Trachinotus ovatus populations within the South China Sea aquaculture context, with the goal of strengthening sustainability. Additionally, the study also enhanced breeding strategies by developing a high-quality haplotype reference panel and an efficient imputation process. Through high-throughput sequencing (depth＞20X), 173 samples from three key breeding sites (SanYa, Hainan; Zhanjiang, Guangdong; and Beihai, Guangxi) were analyzed, revealing substantial genetic diversity and distinct population structures. Notably, the Beihai (BH) population exhibited significant stratification, suggesting regional genetic uniqueness. Low levels of genomic inbreeding coefficient and the identification of unique SNP loci across populations underscored the genetic richness within these Trachinotus ovatus communities. A major constructed of this work is the creation of the first high-quality haplotype reference panel for the species, demonstrating genotype imputation's viability as a cost-effective alternative for large-scale genetic studies in aquaculture. In particular, a Shapeit reference panel with 5476,550 SNPs was created, excelling in accuracy across various depths, notably achieving up to 95.8 % imputation accuracy at 1X sequencing depth. This high accuracy across populations shows the importance of genetic diversity in the panel. This research not only advances the understanding of Trachinotus ovatus genetic foundations but also provides crucial insights for conservation efforts and genetic analysis. By establishing a robust haplotype reference panel for Trachinotus ovatus helps lower the costs of future genetic research and variation studies, this study lays the foundation for sustainable aquaculture practices and breeding programs.

A Comparison of Two Versions of the CRISPR-Sirius System for the Live-Cell Visualization of the Borders of Topologically Associating Domains.

In recent years, various technologies have emerged for the imaging of chromatin loci in living cells via catalytically inactive Cas9 (dCas9). These technologies facilitate a deeper understanding of the mechanisms behind the chromatin dynamics and provide valuable kinetic data that could not have previously been obtained via FISH applied to fixed cells. However, such technologies are relatively complicated, as they involve the expression of several chimeric proteins as well as sgRNAs targeting the visualized loci, a process that entails many technical subtleties. Therefore, the effectiveness in visualizing a specific target locus may be quite low. In this study, we directly compared two versions of a previously published CRISPR-Sirius method based on the use of sgRNAs containing eight MS2 or PP7 stem loops and the expression of MCP or PCP fused to fluorescent proteins. We assessed the visualization efficiency for several unique genomic loci by comparing the two approaches in delivering sgRNA genes (transient transfection and lentiviral transduction), as well as two CRISPR-Sirius versions (with PCP and with MCP). The efficiency of visualization varied among the loci, and not all loci could be visualized. However, the MCP-sfGFP version provided more efficient visualization in terms of the number of cells with signals than PCP-sfGFP for all tested loci. We also showed that lentiviral transduction was more efficient in locus imaging than transient transfection for both CRISPR-Sirius systems. Most of the target loci in our study were located at the borders of topologically associating domains, and we defined a set of TAD borders that could be effectively visualized using the MCP-sfGFP version of the CRISPR-Sirius system. Altogether, our study validates the use of the CRISPR-Sirius technology for live-cell visualization and highlights various technical details that should be considered when using this method.

Mapping taste and flavour traits to genetic markers in lettuce Lactuca sativa

Lettuce is the most highly consumed raw leafy vegetable crop eaten worldwide, making it nutritionally important in spite of its comparatively low nutrient density in relation to other vegetables. However, the perception of bitterness caused by high levels of sesquiterpenoid lactones and comparatively low levels of sweet tasting sugars limits palatability. To assess variation in nutritional and taste-related metabolites we assessed 104 members of a Lactuca sativa cv. Salinas x L. serriola (accession UC96US23) mapping population. Plants were grown in three distinct environments, and untargeted NMR and HPLC were used as a rapid chemotyping method, from which 63 unique Quantitative Trait Loci (QTL) were identified. We were able to identify putative regulatory candidate genes underlying the QTL for fructose on linkage group 9, which accounted for up to 36 % of our population variation, and which was stable across all three growing environments; and for 15-p-hydroxyyphenylacetyllactucin-8-sulfate on linkage group 5 which has previously been identified for its low bitterness, while retaining anti-herbivory field effects. We also identified a candidate gene for flavonoid 3′,5′- hydroxylase underlying a polyphenol QTL on linkage group 5, and two further candidate genes in sugar biosynthesis on linkage groups 2 and 5. Collectively these candidate genes and their associated markers can inform a route for plant breeders to improve the palatability and nutritional value of lettuce in their breeding programmes.

Genetic influences on brain and cognitive health and their interactions with cardiovascular conditions and depression

Approximately 40% of dementia cases could be prevented or delayed by modifiable risk factors related to lifestyle and environment. These risk factors, such as depression and vascular disease, do not affect all individuals in the same way, likely due to inter-individual differences in genetics. However, the precise nature of how genetic risk profiles interact with modifiable risk factors to affect brain health is poorly understood. Here we combine multiple data resources, including genotyping and postmortem gene expression, to map the genetic landscape of brain structure and identify 367 loci associated with cortical thickness and 13 loci associated with white matter hyperintensities (P < 5×10−8), with several loci also showing a significant association with cognitive function. We show that among 220 unique genetic loci associated with cortical thickness in our genome-wide association studies (GWAS), 95 also showed evidence of interaction with depression or cardiovascular conditions. Polygenic risk scores based on our GWAS of inferior frontal thickness also interacted with hypertension in predicting executive function in the Canadian Longitudinal Study on Aging. These findings advance our understanding of the genetic underpinning of brain structure and show that genetic risk for brain and cognitive health is in part moderated by treatable mid-life factors.

Rising Incidence of Inflammatory Bowel Disease in the Asian Subcontinent—An Exploration of Causative Factors

Inflammatory bowel disease (IBD), comprising ulcerative colitis (UC) and Crohn’s disease (CD), is a global health problem with a worldwide increase in incidence. While previously it was more prevalent in Western countries, especially in North America and Europe, there has been a recent sizeable increase in the incidence of IBD in Asia. This article is a brief overview of the causative factors that contribute to this rise in IBD cases in Asian countries. There is a notable disparity between the Asian IBD population and the Western one in terms of age distribution and gender predilection, and genetic studies reveal both shared and unique susceptibility loci. Changes in environmental factors, including alterations in diet, hygiene, and lifestyle, are thought to play a key role in the epidemiological transition observed in Asia. It is crucial to comprehend how genetic predisposition interacts with environmental factors in order to understand the causes of IBD in the Asian population and develop precise interventions to alleviate its impact.

Identifying Regions of the Genome Associated with Conception Rate to the First Service in Holstein Heifers Bred by Artificial Insemination and as Embryo Transfer Recipients.

Heifer conception rate to the first service (HCR1) is defined as the number of heifers that become pregnant to the first breeding service compared to the heifers bred. This study aimed to identify loci associated and gene sets enriched for HCR1 for heifers that were bred by artificial insemination (AI, n = 2829) or were embryo transfer (ET, n = 2086) recipients, by completing a genome-wide association analysis and gene set enrichment analysis using SNP data (GSEA-SNP). Three unique loci, containing four positional candidate genes, were associated (p < 1 × 10-5) with HCR1 for ET recipients, while the GSEA-SNP identified four gene sets (NES ≥ 3) and sixty-two leading edge genes (LEGs) enriched for HCR1. While no loci were associated with HCR1 bred by AI, one gene set and twelve LEGs were enriched (NES ≥ 3) for HCR1 with the GSEA-SNP. This included one gene (PKD2) shared between HCR1 AI and ET services. Identifying loci associated or enriched for HCR1 provides an opportunity to use them as genomic selection tools to facilitate the selection of cattle with higher reproductive efficiency, and to better understand embryonic loss.

Single-cell and genome-wide Mendelian randomization identifies causative genes for gout

BackgroundGout is a prevalent manifestation of metabolic osteoarthritis induced by elevated blood uric acid levels. The purpose of this study was to investigate the mechanisms of gene expression regulation in gout disease and elucidate its pathogenesis.MethodsThe study integrated gout genome-wide association study (GWAS) data, single-cell transcriptomics (scRNA-seq), expression quantitative trait loci (eQTL), and methylation quantitative trait loci (mQTL) data for analysis, and utilized two-sample Mendelian randomization study to comprehend the causal relationship between proteins and gout.ResultsWe identified 17 association signals for gout at unique genetic loci, including four genes related by protein-protein interaction network (PPI) analysis: TRIM46, THBS3, MTX1, and KRTCAP2. Additionally, we discerned 22 methylation sites in relation to gout. The study also found that genes such as TRIM46, MAP3K11, KRTCAP2, and TM7SF2 could potentially elevate the risk of gout. Through a Mendelian randomization (MR) analysis, we identified three proteins causally associated with gout: ADH1B, BMP1, and HIST1H3A.ConclusionAccording to our findings, gout is linked with the expression and function of particular genes and proteins. These genes and proteins have the potential to function as novel diagnostic and therapeutic targets for gout. These discoveries shed new light on the pathological mechanisms of gout and clear the way for future research on this condition.

Identification of novel genes associated with herbicide tolerance in Lentil (Lens culinaris ssp. culinaris Medik.)

Weeds pose a major constraint in lentil cultivation, leading to decrease farmers’ revenues by reducing the yield and increasing the management costs. The development of herbicide tolerant cultivars is essential to increase lentil yield. Even though herbicide tolerant lines have been identified in lentils, breeding efforts are still limited and lack proper validation. Marker assisted selection (MAS) can increase selection accuracy at early generations. Total 292 lentil accessions were evaluated under different dosages of two herbicides, metribuzin and imazethapyr, during two seasons at Marchouch, Morocco and Terbol, Lebanon. Highly significant differences among accessions were observed for days to flowering (DF) and maturity (DM), plant height (PH), biological yield (BY), seed yield (SY), number of pods per plant (NP), as well as the reduction indices (RI) for PH, BY, SY and NP. A total of 10,271 SNPs markers uniformly distributed along the lentil genome were assayed using Multispecies Pulse SNP chip developed at Agriculture Victoria, Melbourne. Meta-GWAS analysis was used to detect marker-trait associations, which detected 125 SNPs markers associated with different traits and clustered in 85 unique quantitative trait loci. These findings provide valuable insights for initiating MAS programs aiming to enhance herbicide tolerance in lentil crop.

Methylation patterns associated with C-reactive protein in racially and ethnically diverse populations

ABSTRACT Systemic low-grade inflammation is a feature of chronic disease. C-reactive protein (CRP) is a common biomarker of inflammation and used as an indicator of disease risk; however, the role of inflammation in disease is not completely understood. Methylation is an epigenetic modification in the DNA which plays a pivotal role in gene expression. In this study we evaluated differential DNA methylation patterns associated with blood CRP level to elucidate biological pathways and genetic regulatory mechanisms to improve the understanding of chronic inflammation. The racially and ethnically diverse participants in this study were included as 50% White, 41% Black or African American, 7% Hispanic or Latino/a, and 2% Native Hawaiian, Asian American, American Indian, or Alaska Native (total n = 13,433) individuals. We replicated 113 CpG sites from 87 unique loci, of which five were novel (CADM3, NALCN, NLRC5, ZNF792, and cg03282312), across a discovery set of 1,150 CpG sites associated with CRP level (p < 1.2E–7). The downstream pathways affected by DNA methylation included the identification of IFI16 and IRF7 CpG-gene transcript pairs which contributed to the innate immune response gene enrichment pathway along with NLRC5, NOD2, and AIM2. Gene enrichment analysis also identified the nuclear factor-kappaB transcription pathway. Using two-sample Mendelian randomization (MR) we inferred methylation at three CpG sites as causal for CRP levels using both White and Black or African American MR instrument variables. Overall, we identified novel CpG sites and gene transcripts that could be valuable in understanding the specific cellular processes and pathogenic mechanisms involved in inflammation.

A genome-wide association study of neutrophil count in individuals associated to an African continental ancestry group facilitates studies of malaria pathogenesis

Background'Benign ethnic neutropenia' (BEN) is a heritable condition characterized by lower neutrophil counts, predominantly observed in individuals of African ancestry, and the genetic basis of BEN remains a subject of extensive research. In this study, we aimed to dissect the genetic architecture underlying neutrophil count variation through a linear-mixed model genome-wide association study (GWAS) in a population of African ancestry (N = 5976). Malaria caused by P. falciparum imposes a tremendous public health burden on people living in sub-Saharan Africa. Individuals living in malaria endemic regions often have a reduced circulating neutrophil count due to BEN, raising the possibility that reduced neutrophil counts modulate severity of malaria in susceptible populations. As a follow-up, we tested this hypothesis by conducting a Mendelian randomization (MR) analysis of neutrophil counts on severe malaria (MalariaGEN, N = 17,056).ResultsWe carried out a GWAS of neutrophil count in individuals associated to an African continental ancestry group within UK Biobank, identifying 73 loci (r2 = 0.1) and 10 index SNPs (GCTA-COJO loci) associated with neutrophil count, including previously unknown rare loci regulating neutrophil count in a non-European population. BOLT-LMM was reliable when conducted in a non-European population, and additional covariates added to the model did not largely alter the results of the top loci or index SNPs. The two-sample bi-directional MR analysis between neutrophil count and severe malaria showed the greatest evidence for an effect between neutrophil count and severe anaemia, although the confidence intervals crossed the null.ConclusionOur GWAS of neutrophil count revealed unique loci present in individuals of African ancestry. We note that a small sample-size reduced our power to identify variants with low allele frequencies and/or low effect sizes in our GWAS. Our work highlights the need for conducting large-scale biobank studies in Africa and for further exploring the link between neutrophils and severe malaria.

Uncovering the architecture of selection in two Bos taurus cattle breeds.

Directional selection alters the genome via hard sweeps, soft sweeps, and polygenic selection. However, mapping polygenic selection is difficult because it does not leave clear signatures on the genome like a selective sweep. In populations with temporally stratified genotypes, the Generation Proxy Selection Mapping (GPSM) method identifies variants associated with generation number (or appropriate proxy) and thus variants undergoing directional allele frequency changes. Here, we use GPSM on two large datasets of beef cattle to detect associations between an animal's generation and 11 million imputed SNPs. Using these datasets with high power and dense mapping resolution, GPSM detected a total of 294 unique loci actively under selection in two cattle breeds. We observed that GPSM has a high power to detect selection in the very recent past (<10 years), even when allele frequency changes are small. Variants identified by GPSM reside in genomic regions associated with known breed-specific selection objectives, such as fertility and maternal ability in Red Angus, and carcass merit and coat color in Simmental. Over 60% of the selected loci reside in or near (<50 kb) annotated genes. Using haplotype-based and composite selective sweep statistics, we identify hundreds of putative selective sweeps that likely occurred earlier in the evolution of these breeds; however, these sweeps have little overlap with recent polygenic selection. This makes GPSM a complementary approach to sweep detection methods when temporal genotype data are available. The selected loci that we identify across methods demonstrate the complex architecture of selection in domesticated cattle.

DNA-methylation variability in normal mucosa: a field cancerization marker in patients with adenomatous polyps.

The phenomenon of field cancerization reflects the transition of normal cells into those predisposed to cancer. Assessing the scope and intensity of this process in the colon may support risk prediction and colorectal cancer prevention. The SWEPIC study, encompassing 1,111 participants for DNA methylation analysis and a subset of 84 for RNA-seq, was employed to detect field cancerization in individuals with adenomatous polyps (AP). Methylation variations were evaluated for their discriminative capability, including in external cohorts, genomic localization, clinical correlations, and associated RNA expression patterns. Normal cecal tissue of individuals harboring an AP in the proximal colon manifested dysregulated DNA methylation compared to tissue from healthy individuals at 558 unique loci. Leveraging these adenoma-related differentially variable and methylated CpGs (aDVMCs), our classifier discerned between healthy and AP-adjacent tissues across SWEPIC datasets (cross-validated ROC AUC [0.63-0.81]), including within age-stratified cohorts. This discriminative capacity was validated in three external sets, differentiating healthy from cancer-adjacent tissue (ROC AUC: [0.82-0.88]). Notably, aDVMC dysregulation correlated with polyp multiplicity. More than 50% of aDVMCs were significantly associated with age. These aDVMCs were enriched in active regions of the genome (p < .001), and associated genes exhibited altered expression in AP-adjacent tissues. Our findings underscore the early onset of field cancerization in the right colon during the neoplastic transformation process. A more extensive validation of aDVMC dysregulation as a stratification tool could pave the way for enhanced surveillance approaches, especially given its linkage to adenoma emergence.

Phenome- and genome-wide analyses of retinal optical coherence tomography images identify links between ocular and systemic health.

The human retina is a multilayered tissue that offers a unique window into systemic health. Optical coherence tomography (OCT) is widely used in eye care and allows the noninvasive, rapid capture of retinal anatomy in exquisite detail. We conducted genotypic and phenotypic analyses of retinal layer thicknesses using macular OCT images from 44,823 UK Biobank participants. We performed OCT layer cross-phenotype association analyses (OCT-XWAS), associating retinal thicknesses with 1866 incident conditions (median 10-year follow-up) and 88 quantitative traits and blood biomarkers. We performed genome-wide association studies (GWASs), identifying inherited genetic markers that influence retinal layer thicknesses and replicated our associations among the LIFE-Adult Study (N = 6313). Last, we performed a comparative analysis of phenome- and genome-wide associations to identify putative causal links between retinal layer thicknesses and both ocular and systemic conditions. Independent associations with incident mortality were detected for thinner photoreceptor segments (PSs) and, separately, ganglion cell complex layers. Phenotypic associations were detected between thinner retinal layers and ocular, neuropsychiatric, cardiometabolic, and pulmonary conditions. A GWAS of retinal layer thicknesses yielded 259 unique loci. Consistency between epidemiologic and genetic associations suggested links between a thinner retinal nerve fiber layer with glaucoma, thinner PS with age-related macular degeneration, and poor cardiometabolic and pulmonary function with a thinner PS. In conclusion, we identified multiple inherited genetic loci and acquired systemic cardio-metabolic-pulmonary conditions associated with thinner retinal layers and identify retinal layers wherein thinning is predictive of future ocular and systemic conditions.

Computationally defined and in vitro validated putative genomic safe harbour loci for transgene expression in human cells.

Selection of the target site is an inherent question for any project aiming for directed transgene integration. Genomic safe harbour (GSH) loci have been proposed as safe sites in the human genome for transgene integration. Although several sites have been characterised for transgene integration in the literature, most of these do not meet criteria set out for a GSH and the limited set that do have not been characterised extensively. Here, we conducted a computational analysis using publicly available data to identify 25 unique putative GSH loci that reside in active chromosomal compartments. We validated stable transgene expression and minimal disruption of the native transcriptome in three GSH sites in vitro using human embryonic stem cells (hESCs) and their differentiated progeny. Furthermore, for easy targeted transgene expression, we have engineered constitutive landing pad expression constructs into the three validated GSH in hESCs.

Dissecting the shared genetic architecture between Alzheimer's disease and frailty: a cross-trait meta-analyses of genome-wide association studies.

Introduction: Frailty is the most common medical condition affecting the aging population, and its prevalence increases in the population aged 65 or more. Frailty is commonly diagnosed using the frailty index (FI) or frailty phenotype (FP) assessments. Observational studies have indicated the association of frailty with Alzheimer's disease (AD). However, the shared genetic and biological mechanism of these comorbidity has not been studied. Methods: To assess the genetic relationship between AD and frailty, we examined it at single nucleotide polymorphism (SNP), gene, and pathway levels. Results: Overall, 16 genome-wide significant loci (15 unique loci) (p meta-analysis < 5 × 10-8) and 22 genes (21 unique genes) were identified between AD and frailty using cross-trait meta-analysis. The 8 shared loci implicated 11 genes: CLRN1-AS1, CRHR1, FERMT2, GRK4, LINC01929, LRFN2, MADD, RP11-368P15.1, RP11-166N6.2, RNA5SP459, and ZNF652 between AD and FI, and 8 shared loci between AD and FFS implicated 11 genes: AFF3, C1QTNF4, CLEC16A, FAM180B, FBXL19, GRK4, LINC01104, MAD1L1, RGS12, ZDHHC5, and ZNF521. The loci 4p16.3 (GRK4) was identified in both meta-analyses. The colocalization analysis supported the results of our meta-analysis in these loci. The gene-based analysis revealed 80 genes between AD and frailty, and 4 genes were initially identified in our meta-analyses: C1QTNF4, CRHR1, MAD1L1, and RGS12. The pathway analysis showed enrichment for lipoprotein particle plasma, amyloid fibril formation, protein kinase regulator, and tau protein binding. Conclusion: Overall, our results provide new insights into the genetics of AD and frailty, suggesting the existence of non-causal shared genetic mechanisms between these conditions.

Detection of Fusarium oxysporum f.sp. lactucae race 1 and 4 via race-specific real-time PCR and target enrichment

Fusarium oxysporum f.sp. lactucae (Fol) causes a vascular disease in lettuce that results in significant yield losses. Race-specific and sensitive real-time PCR assays were developed for Fol races 1 and 4, which are prevalent in Europe. Using genotyping-by-sequencing, unique DNA loci specific to each race were identified and subsequently used for the design of primers and hydrolysis probes. Two assays per race were developed to ensure specificity. The two assays of each race could be run in duplex format, while still giving a sensitivity of 100 fg genomic DNA for all assays. Sample preparation methods were developed for plant tissue, soil, and surfaces, with an extra enrichment step when additional sensitivity was required. By controlling the incubation conditions during the enrichment step, the real-time PCR signal could be matched to the number of spore equivalents in the original sample. When enriching naturally infested soil, down to six conidiospore equivalents L-1 soil could be detected. As enrichment ensures sensitive detection and focuses on living Fol propagules, it facilitates the evaluation of control measures. The developed detection methods for soil and surfaces were applied to samples from commercial lettuce farms and confirmed the prevalence of Fol race 4 in Belgium. Monitoring of soil disinfestation events revealed that despite a dramatic decrease in quantity, the pathogen could still be detected either immediately after sheet steaming or after harvesting the first new crop. The detection method for plant tissue was successfully used to quantify Fol in lettuce inoculated with race 1, race 4 or a combination of both. Under the temperature conditions used, race 4 was more aggressive than race 1, as reflected in larger amounts of DNA of race 4 detected in the roots. These newly developed assays are a promising tool for epidemiological research as well as for the evaluation of control measures.

CRISPR-broad: combined design of multi-targeting gRNAs and broad, multiplex target finding

In CRISPR-Cas and related nuclease-mediated genome editing, target recognition is based on guide RNAs (gRNAs) that are complementary to selected DNA regions. While single site targeting is fundamental for localized genome editing, targeting to expanded and multiple chromosome elements is desirable for various biological applications such as genome mapping and epigenome editing that make use of different fusion proteins with enzymatically dead Cas9. The current gRNA design tools are not suitable for this task, as these are optimized for defining single gRNAs for unique loci. Here, we introduce CRISPR-broad, a standalone, open-source application that defines gRNAs with multiple but specific targets in large continuous or spread regions of the genome, as defined by the user. This ability to identify multi-targeting gRNAs and corresponding multiple targetable regions in genomes is based on a novel aggregate gRNA scoring derived from on-target windows and off-target sites. Applying the new tool to the genomes of two model species, C. elegans and H. sapiens, we verified its efficiency in determining multi-targeting gRNAs and ranking potential target regions optimized for broad targeting. Further, we demonstrated the general usability of CRISPR-broad by cellular mapping of a large human genome element using dCas9 fused to green fluorescent protein.

Genetic architectures of cerebral ventricles and their overlap with neuropsychiatric traits.

The cerebral ventricles are recognized as windows into brain development and disease, yet their genetic architectures, underlying neural mechanisms and utility in maintaining brain health remain elusive. Here we aggregated genetic and neuroimaging data from 61,974 participants (age range, 9 to 98 years) in five cohorts to elucidate the genetic basis of ventricular morphology and examined their overlap with neuropsychiatric traits. Genome-wide association analysis in a discovery sample of 31,880 individuals identified 62 unique loci and 785 candidate genes associated with ventricular morphology. We replicated over 80% of loci in a well-matched cohort of lateral ventricular volume. Gene set analysis revealed enrichment of ventricular-trait-associated genes in biological processes and disease pathogenesis during both early brain development and degeneration. We explored the age-dependent genetic associations in cohorts of different age groups to investigate the possible roles of ventricular-trait-associated loci in neurodevelopmental and neurodegenerative processes. We describe the genetic overlap between ventricular and neuropsychiatric traits through comprehensive integrative approaches under correlative and causal assumptions. We propose the volume of the inferior lateral ventricles as a heritable endophenotype to predict the risk of Alzheimer's disease, which might be a consequence of prodromal Alzheimer's disease. Our study provides an advance in understanding the genetics of the cerebral ventricles and demonstrates the potential utility of ventricular measurements in tracking brain disorders and maintaining brain health across the lifespan.