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Frontotemporal dementia (FTD) is a primary cause of dementia encompassing a broad range of clinical phenotypes and cellular pathologies. Genetic discoveries in FTD have largely been driven by linkage studies in well-documented extended families, explaining most of the patients with a known pathogenic mutation. In the context of complex diseases, it is hypothesized that mutations with reduced penetrance or a combination of low-effect size variants with environmental factors drive disease. Furthermore, these genes are likely to be part of the interaction networks of known FTD genes, contributing to converging cellular processes. In this review, we examine gene discovery approaches in FTD and introduce network biology concepts as tools to assist gene identification studies in genetically complex disease.

Introduction There has been limited research conducted on the identifications/methodological approaches of using plant species as indicators of the presence of economically, important mineral resources. Objectives This study set out to answer the following questions (1) Do specific plant species and species assemblages indicate the presence of mineral deposits? and (2) if yes, then what sort of ecological, experimental, and statistical procedures could be employed to identify such indicators? Methods Keeping in mind these questions, the vegetation of subtropical mineral mines sites in northern Pakistan were evaluated using Indicator Species Analysis (ISA), Canonical Correspondence Analysis (CCA) and Structural Equation Modeling (SEM). Results A total of 105 plant species belonging to 95 genera and 43 families were recorded from the three mining regions. CA and TWCA classified all the stations and plants into three major mining zones, corresponding to the presence of marble, coal, and chromite, based on Jaccard distance and Ward’s linkage methods. This comprehended the following indicator species: Ficus carica, Isodon rugosus and Ajuga parviflora (marble indicators); Olea ferruginea, Gymnosporia royleana and Dicliptera bupleuroides (coal indicators); and Acacia nilotica, Rhazya stricta and Aristida adscensionis (chromite indicators) based on calculated Indicator Values (IV). These indicators were reconfirmed by CCA and SEM analysis. Conclusion It was concluded that ISA is one of the best techniques for the identification/selection of plant indicator species, followed by reconfirmation via CCA and SEM analysis. In addition to establishing a robust approach to identifying plant indicator species, our results could have application in mineral prospecting and detection.

The Hessian fly (HF, Mayetiola destructor) is one of the destructive pests of wheat (Triticum aestivum L.) worldwide. Resistant cultivars can effectively minimize wheat damage due to this insect pest. To identify new quantitative trait loci (QTL) for HF resistance, a population of recombinant inbred lines (RILs) was developed from a cross between the HF-resistant wheat cultivar ‘Chokwang’ and susceptible wheat ‘Ning 7840’, and phenotyped for responses to HF attack. A linkage map was constructed using 1147 single nucleotide polymorphism (SNP) markers generated from genotyping-by-sequencing (GBS). One major QTL, QHf.hwwg-6BS, for HF-resistance was identified on chromosome arm 6BS, which explained up to 84.0% of the phenotypic variation and was contributed by Chokwang. Two RILs showed recombination in the candidate interval of QHf.hwwg-6BS, which delimited QHf.hwwg-6BS to a 4.75 Mb physical interval between 6,028,601 bp and 10,779,424 bp on chromosome arm 6BS of IWGSC Chinese Spring reference genome RefSeq v2.0. Seven GBS-SNPs in the candidate interval were converted into Kompetitive allele specific polymerase chain reaction (KASP) markers. Two of them, KASP-6B112698 and KASP-6B7901215, were validated in a U.S. winter wheat panel. KASP-6B112698 was nearly diagnostic, thus can be used to screen QHf.hwwg-6BS and pyramid it with other resistance genes in breeding programs.

Black point disease caused by Bipolaris sorokiniana is a problem in wheat production worldwide. We aimed to identify major quantitative trait loci (QTL) for resistance to black point and develop molecular markers for marker-assisted selection (MAS). A recombinant inbred line (RIL) population derived from a cross between Wanyuanbai 1 (susceptible) and SN4143 (resistant) was evaluated for black point response at three locations during two years under artificial inoculation with B. sorokiniana, providing data for six environments. Thirty resistant and 30 susceptible RILs were selected to form resistant and susceptible bulks, respectively, that were genotyped by the wheat 660K SNP array; 685 single-nucleotide polymorphisms (SNPs) were identified, among which 385 (56.2%) and 115 (16.8%) were located on chromosomes 4A and 2B, respectively. Bulked segregant RNA-Seq analysis identified candidate regions on chromosomes 4A (4.60 Mb–40.28 Mb) and 5A (1.22 Mb–48.47 Mb). Genetic linkage maps were constructed for chromosomes 2B, 4A, and 5A using 59 polymorphic dCAPS and SSR markers. Finally, two QTL, designated QBB.hau-4A and QBB.hau-5A, were detected on chromosomes 4A and 5A, respectively. The resistance allele of QBB.hau-4A was derived from SN4143, and that of QBB.hau-5A came from Wanyuanbai 1. QBB.hau-4A with a large and consistent effect (15.1%) is likely to be a new locus for black point resistance. The markers linked to QBB.hau-4A and QBB.hau-5A have potential application in MAS-based breeding.

The study of yield traits can reveal the genetic architecture of grain yield for improving maize production. In this study, an association panel comprising 362 inbred lines and a recombinant inbred line population derived from X178 × 9782 were used to identify candidate genes for nine yield traits. High-priority overlap (HPO) genes, which are genes prioritized in a genome-wide association study (GWAS), were investigated using coexpression networks. The GWAS identified 51 environmentally stable SNPs in two environments and 36 pleiotropic SNPs, including three SNPs with both attributes. Seven hotspots containing 41 trait-associated SNPs were identified on six chromosomes by permutation. Pyramiding of superior alleles showed a highly positive effect on all traits, and the phenotypic values of ear diameter and ear weight consistently corresponded with the number of superior alleles in tropical and temperate germplasm. A total of 61 HPO genes were detected after trait-associated SNPs were combined with the coexpression networks. Linkage mapping identified 16 environmentally stable and 16 pleiotropic QTL. Seven SNPs that were located in QTL intervals were assigned as consensus SNPs for the yield traits. Among the candidate genes predicted by our study, some genes were confirmed to function in seed development. The gene Zm00001d016656 encoding a serine/threonine protein kinase was associated with five different traits across multiple environments. Some genes were uniquely expressed in specific tissues and at certain stages of seed development. These findings will provide genetic information and resources for molecular breeding of maize grain yield.

Gilles de la Tourette syndrome (TS) is a common, childhood-onset psychiatric disorder characterized by persistent motor and vocal tics. It is a heterogeneous disorder in which the phenotypic expression may be affected by environmental factors, such as immune responses. Furthermore, several studies have shown that genetic factors play a vital role in the etiology of TS, as well as its comorbidity with other disorders, including attention deficit hyperactivity disorder, obsessive-compulsive disorder, and autism spectrum disorder. TS has a complex inheritance pattern and, according to various genetic studies, several genes and loci have been correlated with TS. Genome-wide linkage studies have identified Slit and Trk-like 1 (SLITRK1) and histidine decarboxylase (HDC) genes, and candidate gene association studies have extensively investigated the dopamine and serotonin system genes, but there have been no consistent results. Moreover, genome-wide association studies have implicated several genetic loci; however, larger study cohorts are needed to confirm this. Copy number variations, which are polymorphisms in the number of gene copies due to chromosomal deletions or duplications, are considered another significant source of mutations in TS. In the last decade, whole genome/exome sequencing has identified several novel genetic mutations in patients with TS. In conclusion, more studies are needed to reveal the exact mechanisms of underlying TS, which may help to provide more information on the prognosis and therapeutic plans for TS.

Oil and protein content and fatty acid composition are quality traits in peanut. Elucidating the genetic mechanisms underlying these traits may help researchers to obtain improved cultivars by molecular breeding. Whole-genome resequencing of a recombinant inbred population of 318 lines was performed to construct a high-density linkage map and identify QTL for peanut quality. The map, containing 4561 bin markers, covered 2032 cM with a mean marker density of 0.45 cM. A total of 110 QTL for oil and protein content, and fatty acid composition were mapped on the 18 peanut chromosomes. The QTL qA05.1 was detected in four environments and showed a major phenotypic effect on the contents of oil, protein, and six fatty acids. The genomic region spanned by qA05.1, corresponding to a physical interval of approximately 1.5 Mb, contains two SNPs polymorphic between the parents that could cause missense mutations. The two SNP sites were employed as KASP markers and validated using lines with extremely high and low oil contents. These sites may be useful in the marker-assisted breeding of peanut cultivars with high oil contents.

Next-generation sequencing (NGS) technologies have transformed medical genetics. However, short-read lengths pose a limitation on identification of structural variants, sequencing repetitive regions, phasing of distant nucleotide changes, and distinguishing highly homologous genomic regions. Long-read sequencing technologies may offer improvements in the characterization of genes that are currently difficult to assess. We used a combination of targeted DNA capture, long-read sequencing, and a customized bioinformatics pipeline to fully assemble the RH region, which harbors variation relevant to red cell donor-recipient mismatch, particularly among patients with sickle cell disease. RHD and RHCE are a pair of duplicated genes located within an ∼175 kb region on human chromosome 1 that have high sequence similarity and frequent structural variations. To achieve the assembly, we utilized palindrome repeats in PacBio SMRT reads to obtain consensus sequences of 2.1 to 2.9 kb average length with over 99% accuracy. We used these long consensus sequences to identify 771 assembly markers and to phase the RHD-RHCE region with high confidence. The dataset enabled direct linkage between coding and intronic variants, phasing of distant SNPs to determine RHD-RHCE haplotypes, and identification of known and novel structural variations along with the breakpoints. A limiting factor in phasing is the frequency of heterozygous assembly markers and therefore was most successful in samples from African Black individuals with increased heterogeneity at the RH locus. Overall, this approach allows RH genotyping and de novo assembly in an unbiased and comprehensive manner that is necessary to expand application of NGS technology to high-resolution RH typing.

Whole genome sequence from Acinetobacter baumannii isolate Ab-46-1632 reveals a novel KL144 capsular polysaccharide (CPS) biosynthesis gene cluster, which carries genes for d -glucuronic acid ( D -GlcA) and l -rhamnose ( l -Rha) synthesis. The CPS was extracted from Ab-46-1632 and studied by 1H and 13C NMR spectroscopy, including a two-dimensional 1H,13C HMBC experiment and Smith degradation. The CPS was found to have a hexasaccharide repeat unit composed of four l -Rhap residues and one residue each of d -GlcpA and N-acetyl- d -glucosamine ( D -GlcpNAc) consistent with sugar synthesis genes present in KL144. The K144 CPS structure was established and found to be related to those of A. baumannii K55, K74, K85, and K86. A comparison of the corresponding gene clusters to KL144 revealed a number of shared glycosyltransferase genes correlating to shared glycosidic linkages in the structures. One from the enzymes, encoded by only KL144 and KL86, is proposed to be a novel multifunctional rhamnosyltransfaerase likely responsible for synthesis of a shared α- l -Rhap-(1 → 2)-α-L-Rhap-(1 → 3)-L-Rhap trisaccharide fragment in the K144 and K86 structures.

Structural variation can create supergene architectures through tight genomic linkages that maintain traits in favourable combinations. A new study by Sodeland et al. links such supergenes in Atlantic cod with species persistence over millennia, despite the fisheries-induced decline in populations. This links intraspecific supergene diversity to ecological stasis, with significant consequences for ecosystem stability.

Litchi chinensis Sonn is widely cultivated in subtropical regions and has an important economic value. A high-density genetic map is a valuable tool for mapping quantitative trait loci (QTL) and marker-assisted breeding programs. In this study, a single nucleotide polymorphism (SNP)-based high-density linkage map was constructed by a genotyping-by-sequencing (GBS) protocol using an F1 population of 178 progenies between two commercial litchi cultivars, ‘Ziniangxi’ (dwarf) and ‘Feizixiao’ (vigorous). The genetic map consisted of 3027 SNP markers with a total length of 1711.97 cM in 15 linkage groups (LGs) and an average marker distance of 0.57 cM. Based on this high-density linkage map and three years of phenotyping, a total of 37 QTLs were detected for eight dwarf-related traits, including length of new branch (LNB), diameter of new branch (DNB), length of common petiole (LCP), diameter of common petiole (DCP), length of internode (LI), length of single leaf (LSL), width of single leaf (WSL), and plant height (PH). These QTLs could explain 8.0 to 14.7% (mean=9.7%) of the phenotypic variation. Among them, several QTL clusters were observed, particularly on LG04 and LG11, which might show enrichment for genes regulating the dwarf-related traits in litchi. There were 126 candidate genes identified within the QTL regions, 55 of which are differentially expressed genes by RNA-seq analysis between ‘Ziniangxi’ and ‘Feizixiao’. These DEGs were found to participate in the regulation of cell development, material transportation, signal transduction, and plant morphogenesis, so they might play important roles in regulating plant dwarf-related traits. The high-density genetic map and QTLs identification related to dwarf traits can provide a valuable genetic resource and a basis for marker-assisted selection and genomic studies of litchi.

Grain number per spikelet (GNS) is a key determinant of grain yield in wheat. A recombinant inbred line population comprising 300 lines was developed from a cross between a high GNS variety H461 and Chinese Spring from which the reference genome assembly of bread wheat was obtained. Both parents and the recombinant inbred lines were genotyped using the wheat 55K single nucleotide polymorphism (SNP) array. A high-density genetic map containing 21,197 SNPs was obtained. These markers covered each of the 21 chromosomes with a total linkage distance of 3792.71 cM. Locations of these markers in this linkage map were highly consistent with their physical locations in the genome assembly of Chinese Spring. The two parents and the whole RIL population were assessed for GNS in two consecutive years at two different locations. Based on multi-environment phenotype data and best liner unbiased prediction values, three quantitative trait loci (QTL) for GNS were identified. One of them located on chromosomes 2B and the other two on 2D. Phenotypic variation explained by these loci varied from 3.07% to 26.57%. One of these QTL, QGns.sicau-2D-2, was identified in each of all trials conducted. Based on the best linear unbiased prediction values, this locus explained 19.59%–26.57% of phenotypic variation. A KASP (Kompetitive Allele-Specific PCR) marker closely linked with this locus was generated and used to validate the effects of this locus in three different genetic backgrounds. The identified QTL and the KASP marker developed for it will be highly valuable in fine-mapping the locus and in exploiting it for marker-assisted selection in wheat breeding programs.

The ratoon stunting disease (RSD) of sugarcane, caused by the bacterium Leifsonia xyli subsp. xyli, is one of the major concerns to sugarcane production and breeding programs worldwide. Due to no obvious external symptoms, RSD cannot be easily detected by the growers, hence has reduced the world’s sugarcane production significantly. This study aimed to identify quantitative trait loci (QTL) associated with RSD resistance and to assist in the development of linked molecular markers for marker-assisted breeding to minimize the reduction in sugarcane yield by the RSD infection. A set of 146 individuals derived from a self-crossing of CP80-1827 were evaluated for RSD resistance in a mechanically duplicated inoculated field trial from 2014 to 2017 using tissue blot immunoassay. Leveraging the genetic data and the four years phenotyping data of CP80-1827 selfing population, linkage map construction and QTL analysis were conducted based on clonal F1 and F2 mapping population types with GACD V.1.2 and IciMapping V.3.3, respectively. A total of 23 QTL associated with RSD resistance were identified, which explained 6% to 13% of the phenotypic variation with the two types of software. A total of 82 disease resistance genes were identified by searching these 23 QTL regions on their corresponding regions on the Sorghum bicolor genome (44 genes), sugarcane R570 genome (20 genes), and S. spontaneum genome (18 genes), respectively. Compared with IciMapping V.3.3, GACD V.1.2 identified more major (6 vs. 3) and stable QTL (2 vs. 0), and more disease resistance genes (51 vs. 31), indicating GACD V.1.2 (clonal F1 mapping type) is most likely to be more efficient than IciMapping (F2 mapping type) for QTL analysis of a sefling population or clonal F1 population in clonal species. The identified QTL controlling RSD resistance along with the associated SNP markers will assist sugarcane molecular breeding programs in combating this disease.

Kernel weight (KW), together with kernel number per unit area, determines yield of cereal crops. Here, two barley recombinant inbred lines (RILs) populations with a shared parent were used to identify loci controlling KW. One is Baudin/AWCS276 (BA) for which a linkage map was available. Several large-effect QTL controlling KW were detected in this population. Another is Morex/AWCS276 (MA). A linkage map with 5273 makers formed 1454 clusters, was constructed by the genotyping by sequence (GBS) data of 201 RILs from this population. A single marker was selected to represent each of the clusters and the linkage map constructed with these markers covers a total length of 1022.4 cM with an average interval of approximately 0.7 cM between loci. Three of the large-effect loci controlling KW (located on 2HL, 6HL, and 7HL, respectively) identified from the BA population were also detected in the MA population under different environments. The locus on 6HL was detected in each of the experiments conducted for both populations thus was selected for developing near isogenic lines (NILs). Apart from KW, the two isolines for each pair of the putative NILs obtained showed no significant difference for any of the morphological characteristics assessed. The average difference in KW between the isolines for all the NILs obtained was about 15% based on assessments under both glasshouse and field conditions. Taken advantage that high quality genome assemblies for both Morex and AWCS276 are available, we identified candidate genes underlying two of the three loci based on an orthologous analysis. The NILs developed and the candidate genes identified in this study should facilitate the cloning and functional analysis of genes regulating KW in barley.

Introduction Membranous nephropathy (MN) is the most common cause of nephrotic syndrome (NS) in adults and is a leading cause of end-stage renal disease due to glomerulonephritis. Primary MN has a strong male predominance, accounting for approximately 65% of cases; yet, currently associated genetic loci are all located on autosomes. Previous reports of familial MN have suggested the existence of a potential X-linked susceptibility locus. Identification of such risk locus may provide clues to the etiology of MN. Methods We identified 3 families with 8 members affected by primary MN. Genotyping was performed using single-nucleotide polymorphism microarrays, and serum was sent for anti-phospholipase A2 receptor (PLA2R) antibody testing. All affected members were male and connected through the maternal line, consistent with X-linked inheritance. Genome-wide multipoint parametric linkage analysis using a model of X-linked recessive inheritance was conducted, and genetic risk scores (GRSs) based on known MN-associated variants were determined. Results Anti-PLA2R testing was negative in all affected family members. Linkage analysis revealed a significant logarithm of the odds score (3.260) on the short arm of the X chromosome at a locus of approximately 11 megabases (Mb). Haplotype reconstruction further uncovered a shared haplotype spanning 2 Mb present in all affected individuals from the 3 families. GRSs in familial MN were significantly lower than in anti-PLA2R–associated MN and were not different from controls. Conclusions Our study identifies linkage of familial membranous nephropathy to chromosome Xp11.3-11.22. Family members affected with MN have a significantly lower GRS than individuals with anti-PLA2R–associated MN, suggesting that X-linked familial MN represents a separate etiologic entity., Graphical abstract

Background Atrial fibrillation (AF) represents the most common clinical cardiac arrhythmia and substantially increases the risk of cerebral stroke, heart failure, and death. Although causative genes for AF have been identified, the genetic determinants for AF remain largely unclear. Objective This study aimed to investigate the molecular basis of AF in a Chinese kindred. Methods A 4-generation family with autosomal-dominant AF and other arrhythmias (atrioventricular block, sinus bradycardia, and premature ventricular contractions) was recruited. Genome-wide scan with microsatellite markers and linkage analysis as well as whole-exome sequencing analysis were performed. Electrophysiological characteristics and subcellular localization of the AF-linked mutant were analyzed using dual whole-cell patch clamps and confocal microscopy, respectively. Results A novel genetic locus for AF was mapped to chromosome 17q21.3, a 3.23-cM interval between markers D17S951 and D17S931, with a maximum 2-point logarithm of odds score of 4.2144 at marker D17S1868. Sequencing analysis revealed a heterozygous mutation in the mapping region, NM_005497.4:c.703A>T;p.(M235L), in the GJC1 gene encoding connexin45 (Cx45). The mutation cosegregated with AF in the family and was absent in 632 control individuals. The mutation decreased the coupling conductance in cell pairs (M235L/M235L, M235L/Cx45, M235L/Cx43, and M235L/Cx40), likely because of impaired subcellular localization. Conclusion This study defines a novel genetic locus for AF on chromosome 17q21.3 and reveals a loss-of-function mutation in GJC1 (Cx45) contributing to AF and other cardiac arrhythmias.

Heavy metal pollutions in the soils are increasingly threatening the global crop and food production. Using plant associated bacteria to remediate heavy metal contamination is a promising approach. We have isolated a cadmium (Cd) resistant Enterococcus faecium strain CX 2–6 from a heavy metal contaminated farmland. We have shown that: (i) CX 2–6 can tolerate cadmium (Cd) with a slower growth rate; (ii) The CX 2–6 complete genome is fully assembled using PacBio long reads; (iii) Differential expression analysis found 47% of CX 2–6 genes are significantly affected by Cd treatment and form three gene groups with distinct expression profiles; (iv) Differentially expressed genes (DEGs) form physically linked gene clusters in the CX 2–6 genome, and one of the gene clusters corresponds to a prophage that is unique to CX 2–6 and is strongly activated when Cd concentration is higher; (v) A majority of DEGs responding to Cd treatment are present in the core genome; and (vi) 55 noncoding RNA genes are identified and 49 of them are DEGs responding to cadmium stress. Our pan-genome analysis and comparative RNA-seq data analysis has significantly improved our understanding of the metabolic reprogramming of E. faecium CX 2–6 under Cd stress.

Graphical abstract, Highlights • The first quantitative synthesis of GWAS regarding Parkinson’s Disease. • Fifteen Parkinson’s Disease GWASs with 191.397 available SNPs pooled. • User friendly software (METRADISC-XL) implemented. • Seven chromosomal regions (bins) were replicated as associated with the Parkinson’s Disease trait., Introduction Parkinson’s disease is a neurodegenerative disorder with a complex etiology coming from interactions between genetic and environmental factors. Research on Parkinson’s disease genetics has been an effortful struggle, while new technologies and novel study designs served as indispensable boosters. Until now, 90 loci and 20 disease-causing gene mutations have been identified. In this study we describe a novel non-parametric approach to GWAS meta-analysis and its application in PD genetics. Methods A literature search was conducted to identify Genome-Wide Association Studies (GWAS) regarding Parkinson’s disease. We applied predefined inclusion criteria and extracted the reported SNPs and their respective position and statistical significance. We divided all chromosomes in approximately equal genetic distance segments called bins and recorded the most significant SNP from each bin and each study and ranked them in terms of their p-value. Ranks from each bin were summed, averaged and added in a heterogeneity-based analysis using the METRADISC-XL software. Weighted and unweighted analysis was performed. Results Five-hundred and forty-three SNPs and their respective p-values from 15 studies were matched in their corresponding bins. The METRADISC-XL analysis resulted in 7 bins with a significant p-value. A bin on chromosome 4 where the SNCA gene is located found with genome-wide significant association with Parkinson’s Disease. Conclusion This is the first time a non-parametric method is applied in GWAS meta-analysis. The results add some insight on the overall understanding of Parkinson’s disease genetics and serve as a first step of further convergent analysis with Genome-wide linkage studies.

Brassica rapa is one of the most important leafy vegetable crops with large cultivated area in China. To increase the availability of DNA markers in B. rapa, we developed insertion-deletion (InDel) markers utilizing high-resolution melting (HRM) curve analysis. We designed primers for 252 InDels (≥ 3 bp) evenly distributed in the genome and tested gene polymorphisms with eight accessions. In total, 208 markers were specifically amplified, and 148 InDels with polymorphism were genotyped successfully using HRM. We further analyzed the correlation with InDel size, GC number, and predicted the difference in Tm values (∆Tm) using 208 markers with specific amplification. We found that the success rate of InDel markers was correlated with the GC number of InDel and the predicted-∆Tm, but not clearly correlated with the length of InDel. When the GC number within InDel was ≥ 8, the successful rate exceeded 90.0%. When the predicted-∆Tm reached 0.5 °C, the success rate was greater than 90.0%, and when it was ≥ 0.6 °C, the rate climbed to 100.0%, indicating their role as the optimal parameter for successful development of an applicable InDel marker. The polymorphic InDel markers can be easily genotyped using HRM. They are of great value in genetic analysis, construction of linkage map, and molecular marker-assisted selection in B. rapa.

The objective of this study was to identify sources and linkages among methicillin-resistant Staphylococcus aureus infections using whole-genome sequencing (WGS).A total of 56 samples were obtained from all patients with a confirmed MRSA infection over 6 months at University of Florida-Health Jacksonville. Samples were cultured and sequenced; data was analyzed on an automated cloud-based platform. Genetic Clusters were defined as40 single nucleotide polymorphisms. Temporal Clusters were defined as ≥5 MRSA cases over 3 days.We found 7 Genetic Clusters comprising 15 samples. Four Genetic Clusters contained patients with non-overlapping stays (3-10 weeks apart), 3 of which contained patients who shared the same Unit. We also found 5 Temporal Clusters comprising 23 samples, although none of the samples were genetically related.Results showed that temporal clustering may be a poor indicator of genetic linkage. Shared epidemiological characteristics between patients in Genetic Clusters may point toward previously unidentified hospital sources. Repeated observation of related strains is also consistent with ongoing MRSA transmission within the surrounding high-risk community.WGS is a valuable tool for hospital infection prevention and control.

Following Hardy-Weinberg disequilibrium (HWD) occurring at a single locus and linkage disequilibrium (LD) between two loci in generations, we here proposed the third genetic disequilibrium in population: recombination disequilibrium (RD). RD is a measurement of crossover interference among multiple loci in a random mating population. In natural populations besides recombination interference, RD may also be due to selection, mutation, gene conversion, drift and/or migration. Therefore, similarly to LD, RD will also reflect the history of natural selection and mutation. In breeding populations, RD purely results from recombination interference and hence can be used to build or evaluate and correct a linkage map. Several practical examples from F2, testcross and human populations indeed demonstrate that RD is useful for measuring recombination interference between two short intervals and evaluating linkage maps. As with LD, RD will be important for studying genetic mapping, association of haplotypes with disease, plant breading and population history.

Background Primary coenzyme Q10 deficiency refers to a group of diseases characterised by reduced levels of coenzyme Q10 in related tissues or cultured cells associated with the 9 genes involved in the biosynthesis of coenzyme Q10. A biallelic pathogenic variant of COQ8A gene causes the occurrence of the primary coenzyme Q10 deficiency type 4. The objective of this study was to investigate the genetic cause of muscle weakness in a proband who had a negative DMD gene test for Becker muscular dystrophy. Methods The DNA of the proband was sequenced using whole exome sequencing. With the help of the Human Phenotype Ontology (HPO), the range of related candidate pathogenic genes has been reduced to a certain extent based on “muscle weakness” (HP:0001324). In addition, family linkage analysis, phenotypic-genotype check and protein structure modeling were used to explore the genetic cause of the proband. Results The compound heterozygous variant c.836A > C (p.Gln279Pro) and c.1228C > T (p.Arg410Ter) in the COQ8A gene was identified in the proband. According to the 2015 American College of Medical Genetics and Genomics (ACMG) standards and guidelines for the interpretation of sequence variants, the novel variant c.836A > C could be classified as “likely pathogenic” for the proband. Conclusion The p.Gln279Pro was detected in the KxGQ motif and the QKE triplet of the COQ8A protein, whose structures were crucial for the structure and function of the COQ8A protein associated with the biosynthesis of coenzyme Q10 and the proband's clinical symptoms were relatively milder than those previously reported.

The first experimental crosses carried out with the human malaria parasite Plasmodium falciparum played a key role in determining the genetic loci responsible for drug resistance, virulence, invasion, growth rate, and transmission. These crosses relied on splenectomized chimpanzees to complete the liver stage of the parasite's life cycle and the subsequent transition to asexual blood stage culture followed by cloning of recombinant progeny in vitro. Crosses can now be routinely carried out using human-liver-chimeric mice infused with human erythrocytes to generate hundreds of unique recombinant progeny for genetic linkage mapping, bulk segregant analysis, and high-throughput 'omics readouts. The high number of recombinant progeny should allow for unprecedented power and efficiency in the execution of a systems genetics approach to study P. falciparum biology.

Congenital hearing loss is a common disorder worldwide. Heterogeneous gene variation accounts for approximately 20-25% of such patients. We investigated a five-generation Chinese family with autosomal-dominant nonsyndromic sensorineural hearing loss (SNHL). No wave was detected in the pure-tone audiometry, and the auditory brainstem response was absent in all patients. Computed tomography of the patients, as well as of two sporadic SNHL cases, showed bilateral inner ear anomaly, cochlear maldevelopment, absence of the osseous spiral lamina, and an enlarged vestibular aqueduct. Such findings were absent in nonaffected persons. We used linkage analysis and exome sequencing and uncovered a heterozygous missense mutation in the PI4KB gene (p.Gln121Arg) encoding phosphatidylinositol 4-kinase β (PI4KB) from the patients in this family. In addition, 3 missense PI4KB (p.Val434Gly, p.Glu667Lys, and p.Met739Arg) mutations were identified in five patients with nonsyndromic SNHL from 57 sporadic cases. No such mutations were present within 600 Chinese controls, the 1000 genome project, gnomAD, or similar databases. Depleting pi4kb mRNA expression in zebrafish caused inner ear abnormalities and audiosensory impairment, mimicking the patient phenotypes. Moreover, overexpression of 4 human missense PI4KB mutant mRNAs in zebrafish embryos resulted in impaired hearing function, suggesting dominant-negative effects. Taken together, our results reveal that PI4KB mutations can cause SNHL and inner ear malformation. PI4KB should be included in neonatal deafness screening.

Chicken plumage color is an important economical trait in poultry breeding, as triple-yellow indigenous broilers are preferred over western commercial broilers in the Chinese market. However, the studies on the pigmentation of plumage coloration are relatively rare at present. Here, we performed a genome-wide mapping study on an F2 intercross, whose 2 founders were one hybrid commercial line “High Quality chicken Line A” that originated from the Anak red chicken and one indigenous line “Huiyang Beard” chicken that is a classical “triple-yellow” Chinese indigenous breed. Moreover, we used an automatic colorimeter that can quantitatively assess the colorations in L∗, a∗, and b∗ values. One major quantitative trait locus (QTL) on chromosome 2 was thus identified by both genome-wide association and linkage analyses, which could explain 10 to 20% of the total phenotypic variance of the b∗ measurements of the back plumage color. Using linkage analysis, 2 additional QTL on chromosome 1 and 20 were also found to be significantly associated with the plumage coloration in this cross. With additional samples from Anak red and Huiyang Beard chickens as well as pooled resequencing data from the 2 founders of this cross, we then further narrowed down the QTL regions and identified several candidate genes, such as CABLES1, CHST11, BCL2L1, and CHD22. As the effects of QTL found in this study were substantial, quantitatively measuring the coloration rather than the descriptive measurements provides stronger statistical power for the analyses. In addition, this major QTL on chromosome 2 that was associated with feather pigmentation at the genome-wide level will facilitate the future chicken breeding for yellow plumage color. In conclusions, we mapped 3 associated QTL on chromosome 1, 2, and 20. The candidate genes identified in this study shed light in the genetic basis of yellow plumage color in chicken.

Oculopharyngodistal myopathy (OPDM) is an adult-onset inherited neuromuscular disorder characterized by progressive ptosis, external ophthalmoplegia, and weakness of the masseter, facial, pharyngeal, and distal limb muscles. The myopathological features are presence of rimmed vacuoles (RVs) in the muscle fibers and myopathic changes of differing severity. Inheritance is variable, with either putative autosomal-dominant or autosomal-recessive pattern. Here, using a comprehensive strategy combining whole-genome sequencing (WGS), long-read whole-genome sequencing (LRS), linkage analysis, repeat-primed polymerase chain reaction (RP-PCR), and fluorescence amplicon length analysis polymerase chain reaction (AL-PCR), we identified an abnormal GGC repeat expansion in the 5′ UTR of GIPC1 in one out of four families and three sporadic case subjects from a Chinese OPDM cohort. Expanded GGC repeats were further confirmed as the cause of OPDM in an additional 2 out of 4 families and 6 out of 13 sporadic Chinese individuals with OPDM, as well as 7 out of 194 unrelated Japanese individuals with OPDM. Methylation, qRT-PCR, and western blot analysis indicated that GIPC1 mRNA levels were increased while protein levels were unaltered in OPDM-affected individuals. RNA sequencing indicated p53 signaling, vascular smooth muscle contraction, ubiquitin-mediated proteolysis, and ribosome pathways were involved in the pathogenic mechanisms of OPDM-affected individuals with GGC repeat expansion in GIPC1. This study provides further evidence that OPDM is associated with GGC repeat expansions in distinct genes and highly suggests that expanded GGC repeat units are essential in the pathogenesis of OPDM, regardless of the genes in which the expanded repeats are located.

BACKGROUND: Inversions involving intron 22 (Inv22) of F8 are detected in approximately 45% of all severe hemophilia A patients. Diagnosis is complicated by the large size of the ~9.5 kb int22h repeated sequence which generates the inversions. Methods such as long-range PCR and inverse-shifting PCR are currently used diagnostically, but suffer from low PCR efficiencies and are difficult to standardize.OBJECTIVES: To design and validate a sensitive and robust assay for the detection of F8 int22h inversions.METHODS: Digital droplet PCR using mile-post assays was used to investigate archival DNA samples.RESULTS: The detection of linkage as a function of physical distance between loci was investigated using an anchor locus and mile-post loci located at 1, 6, 12 and 15 kb distances from the anchor locus. The proportion of linked molecules decreased with increasing distance between loci and showed 30-40% linked molecules for loci 12-15 kb apart. Mile-post assays specific for wild type and Inv22 type 1 and 2 chromosomes were then designed and optimized. All three assays showed high specificities and sensitivities, with coefficients of variation < 5% for all assays. Analysis of 106 patients and 20 carrier mothers showed complete concordance with previously known mutation status. The analysis demonstrated the robustness of the assays versus input DNA concentration (6 ng and higher) and level of fragmentation.CONCLUSIONS: Digital droplet PCR and mile-post assays can be used to detect F8 int22h inversions. The assay systems are technically simple to perform, highly efficient and robust. (Less)

Resistance to powdery mildew is an important trait of interest in many wheat breeding programs. The information on genes conferring resistance to powdery mildew in wheat cultivars is useful in parental selection. Winter wheat breeding line DH51302 derived from Liangxing 99 and cultivar Shimai 26 derived from Jimai 22 showed identical infection patterns against 13 isolates of Blumeria graminis f. sp. tritici (Bgt) that causes wheat powdery mildew. DH51302 and Shimai 26 were crossed to a powdery mildew susceptible cultivar Zhongzuo 9504 and the F2:3 families were used in molecular localization of the resistance genes. Fourteen polymorphic markers, which were linked to Pm52 from Liangxing 99, were used to establish the genetic linkage maps for the resistance genes PmDH51302 and PmSM26 in DH51302 and Shimai 26, respectively. These genes were placed in the same genetic interval where Pm52 resides. Analysis of gene-linked molecular markers indicated that PmDH51302 and PmSM26 differed from other powdery mildew resistance genes on chromosome arm 2BL, such as Pm6, Pm33, Pm51, MlZec1, MlAB10, and Pm64. Based on the results of reaction patterns to different Bgt isolates and molecular marker localization, together with the pedigree information, DH51302 and Shimai 26 carried the same gene, Pm52, which confers their resistance to powdery mildew.

Recombination is a crucial component of evolution and breeding. New combinations of variation on chromosomes are shaped by recombination. Recombination is also involved in chromosomal rearrangements. However, recombination rates vary tremendously among chromosome segments. Genome-wide genetic maps are one of the best tools to study variation of recombination. Here, we describe high density genetic maps of Glycine max and Glycine soja constructed from four segregating populations. The maps were used to identify chromosomal rearrangements and find the highly predictable pattern of cross-overs on the broad scale in soybean. Markers on these genetic maps were used to evaluate assembly quality of the current soybean reference genome sequence. We find a strong inversion candidate larger than 3 Mb based on patterns of cross-overs. We also identify quantitative trait loci (QTL) that control number of cross-overs. This study provides fundamental insights relevant to practical strategy for breeding programs and for pan-genome researches.

This review focuses on gene discovery strategies used to identify monogenic forms of diabetes caused by reduced pancreatic beta-cell number (due to destruction or defective development) or impaired beta-cell function. Gene discovery efforts in monogenic diabetes have identified 36 genes so far. These genetic causes have been identified using four main approaches: linkage analysis, candidate gene sequencing and most recently, exome and genome sequencing. The advent of next-generation sequencing has allowed researchers to move away from linkage analysis (relying on large pedigrees and/or multiple families with the same genetic condition) and candidate gene (relying on previous knowledge on the gene's role) strategies to use a gene agnostic approach, utilizing genetic evidence (such as variant frequency, predicted variant effect on protein function, and predicted mode of inheritance) to identify the causative mutation. This approach led to the identification of seven novel genetic causes of monogenic diabetes, six by exome sequencing and one by genome sequencing. In many of these cases, the disease-causing gene was not known to be important for beta-cell function prior to the gene discovery study. These novel findings highlight a new role for gene discovery studies in furthering our understanding of beta-cell function and dysfunction in diabetes. While many gene discovery studies in the past were led by knowledge in the field (through the candidate gene strategy), now they often lead the scientific advances in the field by identifying new important biological players to be further characterized by in vitro and in vivo studies.

Schizophrenia is a clinically and genetically heterogeneous neuropsychiatric disorder, with a polygenic basis but identification of the specific determinants is a continuing challenge. In this study, we analyzed a multigenerational family, with all healthy individuals in the first two generations, and four progeny affected with schizophrenia in the subsequent two generations, using whole exome sequencing. We identified five rare protein sequence altering heterozygous variants, in five different genes namely SMARCA5, PDE1B, TNIK, SMARCA2 and FLRT shared among all affected members and predicted to be damaging. Variants in SMARCA5 and PDE1B were inherited from the unaffected father whereas variants in TNIK, SMARCA2 and FLRT1 were inherited from the unaffected mother in all the three affected individuals in the third generation; and notably all these five variants were transmitted by an affected mother to her affected son. Microsatellite based analysis lent a modest linkage support (LOD score of 1.2; θ=0.0 at each variant). Of note, analysis of exome data of an ancestry matched unrelated schizophrenia cohort (n = 350), revealed a total of 16 rare variants (MAF 0.01) in these five genes. Interestingly, these five genes involved in neurodevelopmental and/or neurotransmitter signaling processes are implicated in the etiology of schizophrenia previously. This study provides good evidence for a likely cumulative contribution of multiple rare variants from disease relevant genes with a threshold effect in disease development and seems to explain the unusual disease transmission pattern generally witnessed in such conditions, but warrants extensive replication efforts in families with similar complex disease inheritance profiles.

To determine the genetic etiology of deafness in a family (HN-SD01) with autosomal dominant nonsyndromic hearing loss (NSHL). Stepwise genetic analysis was performed on family HN-SD01, including hotspot variant screening, exome sequencing, virtual hearing loss gene panel, and genome-wide linkage analysis. Targeted region sequencing was used to screen ABCC1 in additional cases. Cochlear expression of Abcc1 was evaluated by messenger RNA (mRNA) and protein levels. Computational prediction, immunofluorescence, real-time quantitative polymerase chain reaction, and flow cytometry were conducted to uncover functional consequences of candidate variants. Stepwise genetic analysis identified a heterozygous missense variant, ABCC1:c.1769A>G (p.Asn590Ser), cosegregating with phenotype in HN-SD01. Screening of ABCC1 in an additional 217 cases identified candidate pathogenic variants c.692G>A (p.Gly231Asp) in a sporadic case and c.887A>T (p.Glu296Val) in a familial proband. Abcc1 expressed in stria vascularis and auditory nerve of mouse cochlea. Immunofluorescence showed p.Asn590Ser distributed in cytomembrane and cytoplasm, while wild type was shown only in cytomembrane. Besides, it generated unstable mRNA and decreased efflux capacity of ABCC1. Stepwise genetic analysis is efficient to analyze the genetic etiology of NSHL. Variants in ABCC1 are linked with NSHL and suggest an important role of extruding pumps in maintaining cochlea function.

Average arterial oxyhemoglobin saturation during sleep (AvSpO(2)S) is a clinically relevant measure of physiological stress associated with sleep-disordered breathing, and this measure predicts incident cardiovascular disease and mortality. Using high-depth whole-genome sequencing data from the National Heart, Lung, and Blood Institute (NHLBI) Trans-Omics for Precision Medicine (TOPMed) project and focusing on genes with linkage evidence on chromosome 8p23,1, 2 we observed that six coding and 51 noncoding variants in a gene that encodes the GTPase-activating protein (DLC1) are significantly associated with AvSpO(2)S and replicated in independent subjects. The combined DLC1 association evidence of discovery and replication cohorts reaches genome-wide significance in European Americans (p = 7.9 × 10(−7)). A risk score for these variants, built on an independent dataset, explains 0.97% of the AvSpO(2)S variation and contributes to the linkage evidence. The 51 noncoding variants are enriched in regulatory features in a human lung fibroblast cell line and contribute to DLC1 expression variation. Mendelian randomization analysis using these variants indicates a significant causal effect of DLC1 expression in fibroblasts on AvSpO(2)S. Multiple sources of information, including genetic variants, gene expression, and methylation, consistently suggest that DLC1 is a gene associated with AvSpO(2)S.

Background Syndactyly type II (synpolydactyly, SPD) is a rare autosomal dominant inherited disease with higher incomplete penetrance. Currently, several variants in HOXD13 and one deletion in FBLN1 have been associated with SPD. However, the causative variants in several SPD families and their etiological mechanism are still largely unknown. Methods Whole exome and PCR-sanger sequencing followed by two-point linkage analysis were performed to identify the pathogenic variant in a six-generation Chinese pedigree. Homology modeling in combination with the RNAi and qRT-PCR experiments was used for revealing the pathogenic mechanism of the TTC30B variant. Results A six-generation SPD family was reported. The affected subjects in this family had no other clinical malformation beyond SPD. A rare missense variant c.1157C>T [p.Ala375Val] (chr2:178416368, hg19) in TTC30B was demonstrated to be responsible for this SPD family. The modeling structure indicated that the Ala375 was evolutionarily and structurally conserved. The variant p.Ala375Val was predicted to be deleterious for protein structure and/or stability. Two-point linkage analysis resulted in a maximum LOD score of 3.1444 (P = 0.000071). Furthermore, we found that TTC30B was regulated by the Shh signaling pathway and the abnormal expression of TTC30B will affect the activation of the Shh signaling pathway in human retinal pigment epithelial cells. Conclusions This study demonstrates for the first time that an IFT (intraflagellar transport) - related gene TTC30B is implicated with SPD.

Ecological interactions are largely determined by adaptive traits deemed 'accessory'. In plants, fungi, and bacteria, such traits mainly comprise metabolic pathways that produce or transform diverse molecules. While accessory metabolic pathways are pervasive, it is often difficult to identify their genetic bases. Recently, in-depth descriptions of metabolic gene clusters (MGCs), which encode discrete metabolic pathways, have greatly simplified the characterization of genotype-phenotype maps, yet questions of how this genome architecture relates to the evolution of accessory functions remain. Fungi are uniquely positioned to spearhead investigations into these dynamics because they display gradients in clustering across pathways and taxa. This review will focus on the role of MGCs as both agents and consequences of the accessory function evolution that underpins fungal diversification.

To analyze family-based whole-genome sequence (WGS) data for complex traits, we developed a rare variant (RV) non-parametric linkage (NPL) analysis method, which has advantages over association methods. The RV-NPL differs from the NPL in that RVs are analyzed, and allele sharing among affected relative-pairs is estimated only for minor alleles. Analyzing families can increase power because causal variants with familial aggregation usually have larger effect sizes than those underlying sporadic diseases. Differing from association analysis, for NPL only affected individuals are analyzed, which can increase power, since unaffected family members can be susceptibility variant carriers. RV-NPL is robust to population substructure and admixture, inclusion of nonpathogenic variants, as well as allelic and locus heterogeneity and can readily be applied outside of coding regions. In contrast to analyzing common variants using NPL, where loci localize to large genomic regions (e.g., >50 Mb), mapped regions are well defined for RV-NPL. Using simulation studies, we demonstrate that RV-NPL is substantially more powerful than applying traditional NPL methods to analyze RVs. The RV-NPL was applied to analyze 107 late-onset Alzheimer disease (LOAD) pedigrees of Caribbean Hispanic and European ancestry with WGS data, and statistically significant linkage (LOD ≥ 3.8) was found with RVs in PSMF1 and PTPN21 which have been shown to be involved in LOAD etiology. Additionally, nominally significant linkage was observed with RVs in ABCA7, ACE, EPHA1, and SORL1, genes that were previously reported to be associated with LOAD. RV-NPL is an ideal method to elucidate the genetic etiology of complex familial diseases.

Objective: Duchenne Muscular Dystrophy is an X-linked recessive disorder characterized by progressive muscular degeneration, patients often develop cardiac failure in the later stage and death occurs before 20 years of age. For a disease with poor postnatal prognosis such as Duchenne Muscular Dystrophy (DMD), providing the carrier mother with the option of prenatal diagnosis in a subsequent pregnancy is accepted practice in many places where termination of pregnancy is allowed. Though methods of direct sequencing such as Sanger's sequencing has been widely used, Next-Generation Sequencing is been increasingly replacing most of its application. For the DMD gene, being the longest gene in the human genome, methods of direct sequencing is often unpractical and time-consuming, instead, STR analysis for linkage analysis would be a cost-effective option and have been used routinely for prenatal diagnosis of DMD. The diagnostic significance of the STRs is based on several criteria, the most important one being the heterozygosity of the locus, power of discrimination (PD) and power of exclusion (PE). Material and methods: In this study, we investigated the feasibility of application and diagnostic value of 6 STR loci (DSTR49, DSTR50, DXS1036, DXS1067, DXS890, DXS9907) in the proximity of the DMD gene, 66 healthy individuals were recruited for STR analysis and 5 cases of prenatal diagnosis for carrier mother were performed. Result: Allele frequency, heterozygosity, polymorphic information content, the power of discrimination and exclusion and Hardy–Weinberg equilibrium were analyzed and calculated for the 6 STR loci. 5 of these loci (DSTR49, DSTR50, DXS1067, DXS890, DXS9907) were found practical and useful for preimplantation Genetic diagnosis (PGD) and prenatal diagnosis. All 5 cases of prenatal diagnosis using the method had informative STR results and correct diagnosis. Conclusion: We concluded that our protocol of STR analysis can be applied for prenatal diagnosis and pre-implantation genetic diagnosis of DMD with high confidence and accuracy, especially in clinical settings where diagnostic resources are more limited. Keywords: Duchenne muscular dystrophy, Dystrophin, STR analysis, Prenatal diagnosis

Cluster bean (Cyamopsis tetragonoloba), also known as guar, is an important industrial crop owing to its high gum content in the endosperm. Availability of sufficient genomic resources, especially, DNA markers, greatly aids genetic improvement of a crop. In this study, we identified 1859 genomic SSRs, for the first time, from 1091 scaffolds representing 60% of the cluster bean genome. Further we validated 89 of these markers using 54 cultivated guar accessions and two wild relatives, Cyamopsis serrata and Cyamopsis senegalensis. Seven SSRs were monomorphic even with the wild relatives while 11 were polymorphic only between species with 72 being polymorphic within C. tetragonoloba accessions. Polymorphism information content of the markers ranged from 0.017 to 0.62 with an average of 0.19. Cross-transferability rates of 62% observed for the genomic SSRs suggested divergence between the cultivated and the wild species. Genomic SSRs mined in this study though showed a high proportion of dinucleotide repeats (48.5%), while tri- and tetranucleotide repeats were found to be more polymorphic. Genetic diversity analysis of the 56 accessions using the 82 polymorphic markers could differentiate the cultivated accessions of C. tetragonoloba into four major clusters, two of which had two sub-clusters while the wild accessions formed a separate cluster. Since chromosome-wide distribution of the SSRs is unknown and genetic linkage maps in guar is not available, we used the soybean genome as a reference and identified 29 genome-wide and unlinked SSRs markers. Population structure analysis (PSA) using these markers revealed six subpopulations, more or less similar to the major and sub-clusters identified by the neighbor joining analysis. Further PSA identified an entry from subpopulation 6 to have admixture with the wild relatives. Annotation of the validated genomic SSR containing sequences using green plant nr protein database revealed that 16 of them were genic in nature. This is the first report on genomic SSRs and their utilization in unraveling the genetic diversity in cluster bean.

Relationship estimation and segment detection between individuals is an important aspect of disease gene mapping. Existing methods are either tailored for computational efficiency or require phasing to improve accuracy. We developed TRUFFLE, a method that integrates computational techniques and statistical principles for the identification and visualization of identity-by-descent (IBD) segments using un-phased data. By skipping the haplotype phasing step and, instead, relying on a simpler region-based approach, our method is computationally efficient while maintaining inferential accuracy. In addition, an error model corrects for segment break-ups that occur as a consequence of genotyping errors. TRUFFLE can estimate relatedness for 3.1 million pairs from the 1000 Genomes Project data in a few minutes on a typical laptop computer. Consistent with expectation, we identified only three second cousin or closer pairs across different populations, while commonly used methods identified a large number of such pairs. Similarly, within populations, we identified many fewer related pairs. Compared to methods relying on phased data, TRUFFLE has comparable accuracy but is drastically faster and has fewer broken segments. We also identified specific local genomic regions that are commonly shared within populations, suggesting selection. When applied to pedigree data, we observed 99.6% accuracy in detecting 1(st) to 5(th) degree relationships. As genomic datasets become much larger, TRUFFLE can enable disease gene mapping through implicit shared haplotypes by accurate IBD segment detection.

Pure lines derived from multiple parents provide abundant variation for genetic study. However, efficient genetic analysis methods and user-friendly software are still lacking. In this study, we developed linkage analysis methods and integrated analysis software for pure-line populations derived from four-way and eight-way crosses. First, polymorphic markers are classified into different categories according to the number of identifiable alleles in the inbred parents. Expected genotypic probability is then derived for each pair of complete markers, and based on them a maximum likelihood estimate (MLE) of recombination frequency is calculated. An EM algorithm is proposed for calculating recombination frequencies in scenarios that at least one marker is incomplete. A linkage map can thus be constructed using estimated recombination frequencies. We describe a software package called GAPL for recombination frequency estimation and linkage map construction in multi-parental pure-line populations. Both simulation studies and results from a reported four-way cross recombinant inbred line population demonstrate that the proposed method and software can build more accurate linkage maps in shorter times than other published software packages. The GAPL software is freely available from www.isbreeding.net and can also be used for QTL mapping in multi-parental populations. Keywords: Pure lines, Four-way cross, Eight-way cross, Recombination frequency estimation, Integrated software

Introduction Neurofascin, encoded by NFASC, is a transmembrane protein that plays an essential role in nervous system development and node of Ranvier function. Anti-Neurofascin autoantibodies cause a specific type of chronic inflammatory demyelinating polyneuropathy (CIDP) often characterized by cerebellar ataxia and tremor. Recently, homozygous NFASC mutations were recently associated with a neurodevelopmental disorder in two families. Methods A combined approach of linkage analysis and whole-exome sequencing was performed to find the genetic cause of early-onset cerebellar ataxia and demyelinating neuropathy in two siblings from a consanguineous Italian family. Functional studies were conducted on neurons from induced pluripotent stem cells (iPSCs) generated from the patients. Results Genetic analysis revealed a homozygous p.V1122E mutation in NFASC. This mutation, affecting a highly conserved hydrophobic transmembrane domain residue, led to significant loss of Neurofascin protein in the iPSC-derived neurons of affected siblings. Conclusions The identification of NFASC mutations paves the way for genetic research in the developing field of nodopathies, an emerging pathological entity involving the nodes of Ranvier, which are associated for the first time with a hereditary ataxia syndrome with neuropathy.

We constructed a high-density genetic linkage map of bronze loquat (Eriobotrya deflexa) for a three-way cross of loquat (Eriobotrya japonica) × (loquat × bronze loquat) by using single nucleotide polymorphism (SNP) markers generated by RAD-Seq, with the combination of simple sequence repeat (SSR) markers, which have been used in the previous study. The map of bronze loquat (‘Taiwan loquat No. 1’) consisted of 960 loci including 793 SNPs, 166 SSRs (98 apple, 8 loquat and 60 pear SSRs), and the loquat canker resistance gene Pse-a. This map covered 17 linkage groups over a total length of 1707.4 cM with an average distance of 1.78 cM between markers. The number of linkage groups corresponded to the basic chromosome number (n = 17). The analysis of the markers that can be anchored to apple draft genome showed that almost all loquat linkage groups were aligned to the apple draft genome. The constructed map will enable us to determine the location of quantitative trait loci and genes of interest, and to analyze genome synteny in the tribe Pyreae, subfamily Spiraeoideae of the family Rosaceae.

Many achievements have been made to develop quantitative trait loci (QTLs) and gene-associated single nucleotide polymorphisms (SNPs) to facilitate practical marker-assisted selection (MAS) in aquatic animals. However, the systematic studies of SNPs associated with extreme threshold traits were poor in populations lacking of parental genomic information. Coupling next generation sequencing with bulked segregant analysis (BSA) should allow identification of numerous associated SNPs with extreme phenotypes. In the present study, using combination of SNP frequency difference and Euclidean distance, we conducted linkage analysis of SNPs located in genes involved in immune responses, and identified markers associated with Vibrio anguillarum resistance in turbot (Scophthalmus maximus). A total of 221 SNPs was found as candidate SNPs between resistant and susceptible individuals. Among these SNPs, 35 loci located in immune related genes were genotyped in verification population and 7 of them showed significant association with V. anguillarum resistance in both alleles and genotypes (P 0.05). Among these 7 genes, PIK3CA-like, CYLD, VCAM1, RhoB and RhoGEF are involved in PI3K/Akt/mTOR pathway and NF-κB pathway, which influence the efficiency of bacteria entering the host and inflammation. SNP-SNP interaction analysis was performed by generalized multifactor dimensionality reduction (GMDR). The combination of SNP loci in RhoB, PIK3CA-like and ADCY3 showed a significant effect on V. anguillarum resistance with the verification rate in the sequencing population up to 70.8%. Taken all, our findings demonstrated the feasibility of BSA-seq approach in identifying genes responsible for the extreme phenotypes and will aid in performing MAS in turbot.

The discovery of thermo-sensitive genic male sterility (TGMS) has led to development of a simple and highly efficient two-line breeding system. In this study, genetic analysis was conducted using three F2 populations derived from crosses between IR68301S, an indica TGMS rice line, and IR14632 (tropical japonica), Supanburi 91062 (indica) and IR67966-188-2-2-1 (tropical japonica), respectively. Approximately 1:3 ratio between sterile and normal pollen of F2 plants from the three populations revealed that TGMS is controlled by a single recessive gene. Bulked segregant analysis using simple sequence repeat (SSR) and insertion-deletion (InDel) markers were used to identify markers linked to the tms gene. The linkage analysis based on the three populations indicated that the tms locus was located on chromosome 2 covering the same area. Using IR68301S × IR14632 F2 population, the results showed that the tms locus was located between SSR marker RM12676 and InDel marker 2gAP0050058. The genetic distance from the tms gene to these two flanking markers were 1.10 and 0.82 cM, respectively. InDel marker 2gAP004045 located between these two markers showed complete co-segregation with the TGMS phenotype. In addition, InDel marker vf0206114052 showed 2.94 cM linked to the tms gene using F2 populations of IR68301S × Supanburi 91062. These markers are useful tool for developing new TGMS lines by marker-assisted selection. There were ten genes located between the two flanking markers RM12676 and 2gAP0050058. Using quantitative real-time PCR for expression analysis, 7 of the 10 genes showed expression in panicles, and response to temperatures. These genes could be the candidate gene controlling TGMS in IR68301S. Keywords: hybrid rice, thermo-sensitive genic male sterility, insertion/deletion, simple sequence repeat, marker-assisted selection

Tuber starch content and plant maturity are two important agronomic traits of potato. To investigate the complex genetic basis of these traits in the cultivated potato, as well as the relationship between them, we developed a linkage map in a tetraploid population of 192 clones derived from the cross Longshu 8 × Zaodabai and mapped quantitative trait loci (QTL) for tuber starch content and plant maturity using data collected in three diverse environments over two years. We detected eleven QTL for tuber starch content distributed on seven chromosomes, of which four, on chromosomes I, II, and VIII, were expressed in at least three environments. For plant maturity, we identified six QTL on chromosomes II, IV, V, VII, and XI, one of which, on chromosome V, showed LOD peaks ranging from 45.2 to 62.5 cM and explained 21.6%–26.6% of phenotypic variation was expressed in five of the six environments. Because the reproducible QTL for plant maturity and tuber starch content mapped to different chromosomes and neither overlapping QTL, nor any genetic interaction between QTL were detected, we infer that tuber starch content and plant maturity are controlled by independent genetic loci. This inference supports the prospect of breeding potato for both early maturity and high starch content. Keywords: Potato, QTL, Tetraploid, Tuber starch content, Plant maturity

Host genetic factors are frequently ascribed to differential malaria outcomes as a by-product of evolutionary adaptation. To this respect, Tumor Necrosis factor alpha (TNF-α), a human cytokine, is known to be associated with malaria through its differential regulation in diverse malaria manifestations. Since diversity in differential malaria outcome is uncommon in every endemic settings, possible association of TNF-α and malaria is not commonly established. In order to check for association between the occurrence of Single Nucleotide Polymorphisms (SNPs) in the TNF-α gene with different malaria manifestations, we have sequenced a 4011 bp region constituting the promoter and the whole gene of human TNF-α in 61 patients [(16 cerebral plus severe (SCM), 21 severe (SM) and 24 uncomplicated (UM)] samples in a highly malaria endemic state (Odisha) of India. Multiple sequence alignment revealed presence of six SNPs (−1031 T > C, −863C > A, −857C > T, −308G > A, −806C > T, +787C > A), out of which the -806C > T and +787C > A are novel in malaria patients in general and the +787C > A was detected for the first time in humans. Although alleles due to six different SNPs segregate differentially in the three groups of malaria (SCM, SM and UM) in the present study, interestingly, for the −1031 T > C position, the frequency of individuals possessing the homozygous rare allele was higher in the SCM group with a higher number of heterozygotes in the UM group. The Tajima's D values considering all the SNPs in a defined group were positive and statistically insignificant conforming no evolutionary constraint. However, statistically significant deviation from expectation under Hardy-Weinberg equilibrium for -1031 T > C SNP in the UM group points towards the probable role of natural selection providing some kind of protection to malaria in Odisha, India.

Next generation sequencing technologies have become affordable for most plant breeding programs. In this study we sequenced the entire genome of the Solanum lycopersicum L. cultivar Caimanta and S. pimpinellifolium L. accession LA0722 with assembly relative to the Heinz 1706 reference version SL2.50. We present 1) analysis of the amount and distribution of polymorphism in “Caimanta” and “LA0722” 2) examination of alleles in candidate genes affecting disease resistance, fruit shape, fruit weight and fruit quality and 3) development of molecular markers to construct a genetic linkage map based on a F2 population. A total of 1,397,518 polymorphisms were detected in the comparison between “Caimanta” and “LA0722”. A resistant allele for Rx4/Xv3 was detected by sequence, and confirmed through inoculation. - We developed a set of insertion/deletion (InDel) DNA markers that can be multiplexed and scored using easily accessed genotyping platforms. These markers were used to construct a genetic linkage map. We demonstrate that the whole genome sequencing of parental lines can be successfully used to reveal phenotypes and characterize a reference population through easily accessed genotyping strategies. Fil: Cambiaso, Vladimir. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Investigaciones en Ciencias Agrarias de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Agrarias. Instituto de Investigaciones en Ciencias Agrarias de Rosario; Argentina Fil: Pratta, Guillermo Raúl. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Investigaciones en Ciencias Agrarias de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Agrarias. Instituto de Investigaciones en Ciencias Agrarias de Rosario; Argentina Fil: Pereira Da Costa, Javier Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Investigaciones en Ciencias Agrarias de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Agrarias. Instituto de Investigaciones en Ciencias Agrarias de Rosario; Argentina Fil: Zorzoli, Roxana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Investigaciones en Ciencias Agrarias de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Agrarias. Instituto de Investigaciones en Ciencias Agrarias de Rosario; Argentina Fil: Francis, David Merrill. Ohio State University; Estados Unidos Fil: Rodríguez, Gustavo Rubén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Investigaciones en Ciencias Agrarias de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Agrarias. Instituto de Investigaciones en Ciencias Agrarias de Rosario; Argentina

Several genome-wide association studies (GWAS) identified new single nucleotide polymorphisms (SNPs) with susceptibility to Tuberculosis (TB). However, many of them were not replicated across ethnic groups. The cause of this phenomenon of genetic heterogeneity is uncertain. Here, we attempted to replicate and evaluate the mechanism that causes genetic heterogeneity in several putative TB predisposition loci found by previous GWAS, including chromosome 18q, ASAP1, DUSP14, and HLA-DQA1. A Chinese cohort of 1200 TB patients and 1280 population controls were genotyped. The results showed that genetic predisposition to TB might operate in an age-specific manner. While no significant association was found in the whole samples, a SNP of HLA-DQA1, rs9272785, showed suggestive association within the young-onset TB subgroup (onset at 20–40 years of age, N = 396). The results provide support for the hypothesis that there are different pathogenesis mechanisms causing clinical TB disease in different age groups, and that genetics probably play a substantial role only in young-onset TB.