Polymorphic Single Nucleotide Polymorphism Research Articles

Bulked Segregant Analysis by Sequencing-Based Genetic Mapping of the Green Spotted Fruit Rind Regulating Locus in Wild Melon XNM020 Reveals Four Possible Candidate Genes

Fruit rind patterns are vital commercial quality traits in melon, in which the spotted or striped fruit rinds contribute to the commercial value of melon and can directly affect the choice of consumers. Although the spotted or non-spotted fruit rind pattern was studied in several cultivated melon accessions, the inheritance and regulating locus of this trait in wild melons are still unknown. Therefore, in this study, the inheritance and regulating loci of the green spotted fruit rind in a wild melon accession XNM020 were explored with F2 segregating populations derived from crossing between XNM020 and a cultivated melon XNM125. Segregating ratios of phenotypic data indicated that the green spotted fruit rind in XNM020 has a monogenic dominant inheritance. BSA-Seq showed that two potential genomic regions on chromosomes 4 (from 0.00 to 2.97 Mb) and 5 (from 0.00 to 2.34 Mb) regulate the formation of the green spotted fruit rind in wild melon XNM020. According to the annotations of polymorphic SNPs (single-nucleotide polymorphisms) and small InDels (insertions and deletions) in target genomic regions and the predicted gene functions, four genes MELO3C003316, MELO3C003375, MELO3C003388, and MELO3C014660 regulating chloroplast development or chlorophyll biosynthesis may be the best candidate genes. The results of this study enriched the inheritances of spotted fruit rinds in melon and also provided target genomic regions for marker-assisted selection breeding of melon focusing on fruit rinds.

Association of heat-shock protein 70.1 gene with physiological and physical performance of Bali cattle.

Global warming challenges cattle productivity and welfare since it affects heat stress and scarce feed. The heat-shock protein 70 (HSP70) gene is essential in cytoprotection against stressors, protecting cells from dysregulated gene expression and apoptosis. This study aimed to identify significant genetic markers of the HSP70.1 gene that can be leveraged genetically to enhance thermotolerance and production in Bali cattle further. Animals were sampled from three different rearing systems. In this study, 83 healthy adult male Bali cattle without abnormalities were utilized. Single-nucleotide polymorphism (SNP) diversity associated with the physiological and physical traits of Bali cattle was assessed using SNPStat online software. Gene expression for putative SNPs and their genotypic groups was further evaluated. There were 15 polymorphic SNPs (c.-185G>A, c.-69T>G, c.10G>C, c.19A>G, c.45C>T, c.101INS, c.115T>C, c.130T>C, c.136G>T, c.159G>C, c.164G>T, c.234G>A, c.303G>A, c.333C>A, and c.456C>T) identified, of which 12 were associated with the assessed trait. Nine SNPs were associated with physiological traits, while eight were with physical traits. The c.136G>T as a novel, high minor allele frequency, and associative SNP was selected for HSP70 gene expression. Individuals with the TT genotype have a trim physique, susceptible physiology, and high HSP70 mRNA expression. On the other hand, the GG genotype was significantly associated with larger physique, lower physiology, and low HSP70 mRNA expression. The higher expression may indicate that HSP70.1 is involved in mitigating the deleterious effects of stress. As a result, the animal experienced negative energy balance, decreasing body size. Single-nucleotide polymorphism c.136G>T is a candidate biomarker for heat resistance traits in Bali cattle.

Population structure, runs of homozygosity analysis and construction of single nucleotide polymorphism fingerprinting database of Longnan goat population

AbstractThe Longnan goat (LN) is a local breed discovered during the third census of China's livestock and poultry genetic resources. In order to thoroughly comprehend the population traits, this research examined a total of 70 animals from three different goat populations, namely the LN goat, Nanjiang Brown goat (NJ), and Boer goat (Boer). Specific‐Locus Amplified Fragment Sequencing was employed to analyze genetic diversity, population structure, and selective sweep patterns. Additionally, individual DNA fingerprints were generated to provide detailed genetic profiles for each subject. A total of 643,868 single nucleotide polymorphisms (SNPs) were detected, with a majority located in intergenic and intronic regions. Genetic diversity analysis uncovered lower diversity within the LN population compared to the other two populations. The analysis of population structure unveiled significant genetic distance between LN and both NJ and Boer populations, with distinct clustering patterns observed. Moreover, genetic differentiation coefficients (FST) of 0.1019 and 0.0854 were determined between LN and Boer, as well as LN and NJ, respectively, indicating substantial genetic differentiation. Selective sweep analysis, combining FST and π ratio, identified several genes associated with growth and development, reproduction, hair color, and immunity that may serve as valuable candidates for marker‐assisted breeding. Furthermore, identification of 47,541 runs of homozygosity (ROHs) revealed non‐uniform distribution across chromosomes, with the highest frequency on chromosome 1 and the lowest on chromosome 27. Correlations between different inbreeding coefficients varied, with the highest observed between FHOM and FGRM, and the lowest between FROH >1.5Mb and FROH <500kb. A total of 46 candidate genes were annotated within high‐frequency ROH islands, primarily associated with biological processes such as reproduction, growth and development, and immunity. Finally, a DNA fingerprint, consisting of 371 highly polymorphic SNPs, was generated and presented in the form of a two‐dimensional code for convenient access. Based on the population structure analysis, LN goats have been found to have a distant genetic distance and a higher degree of differentiation from both the Boer and NJ populations. Evaluation of genetic diversity parameters and ROH analysis indicates that the LN population exhibits lower genetic diversity and shows signs of inbreeding. Our findings offer theoretical support for the identification of genetic resources within this population.

Genetic diversity, linkage disequilibrium, and population structure of tetraploid wheat landraces originating from Europe and Asia

BackgroundDurum wheat is one of the most important crops, especially in the Mediterranean region. Insight into the genetic diversity of germplasm can improve the breeding program management in various traits. This study was done using single nucleotide polymorphisms (SNP) markers to characterize the genetic distinctiveness and differentiation of tetraploid wheat landraces collected from nine European and Asian countries. A sum of 23,334 polymorphic SNPs was detected in 126 tetraploid wheat landraces in relation to the reference genome.ResultsThe number of identified SNPs was 11,613 and 11,721 in A and B genomes, respectively. The highest and lowest diversity was on 6B and 6 A chromosomes, respectively. Structure analysis classified the landraces into two distinct subpopulations (K = 2). Evaluating the principal coordinate analysis (PCoA) and weighted pair-group method using arithmetic averages (WPGMA) clustering results demonstrated that landraces (99.2%) are categorized into one of the two chief subpopulations. Therefore, the grouping pattern did not clearly show the presence of a clear pattern of relationships between genetic diversity and their geographical derivation. Part of this result could be due to the historical exchange between different germplasms. Although the result did not separate landraces based on their region of origin, the landraces collected from Iran were classified into the same group and cluster. Analysis of molecular variance (AMOVA) also confirmed the results of population structure. Finally, Durum wheat landraces in some countries, including Turkey, Russia, Ukraine, and Afghanistan, were highly diverse, while others, including Iran and China, were low-diversity.ConclusionThe recent study concluded that the 126 tetraploid wheat genotypes and their GBS-SNP markers are very appropriate for quantitative trait loci (QTLs) mapping and genome-wide association studies (GWAS). The core collection comprises two distinct subpopulations. Subpopulation II genotypes are the most diverse genotypes, and if they possess desired traits, they may be used in future breeding programs. The degree of diversity in the landraces of countries can provide the ground for the improvement of new cultivars with international cooperation. linkage disequilibrium (LD) hotspot distribution across the genome was investigated, which provides useful information about the genomic regions that contain intriguing genes.

Genome-wide association analysis of tan spot disease resistance in durum wheat accessions from Tunisia.

Background: Tunisia harbors a rich collection of unexploited durum wheat landraces (Triticum durum ssp. durum) that have been gradually replaced by elite cultivars since the 1970s. These landraces represent an important potential source for broadening the genetic background of elite durum wheat cultivars and for the introgression of novel genes for key traits, including disease resistance, into these cultivars. Methods: In this study, single nucleotide polymorphism (SNP) markers were used to investigate the genetic diversity and population structure of a core collection of 235 durum wheat accessions consisting mainly of landraces. The high phenotypic and genetic diversity of the fungal pathogen Pyrenophora tritici-repentis (cause of tan spot disease of wheat) in Tunisia allowed the assessment of the accessions for tan spot resistance at the adult plant stage under field conditions over three cropping seasons. A genome-wide association study (GWAS) was performed using a 90k SNP array. Results: Bayesian population structure analysis with 9191 polymorphic SNP markers classified the accessions into two groups, where groups 1 and 2 included 49.79% and 31.49% of the accessions, respectively, while the remaining 18.72% were admixtures. Principal coordinate analysis, the unweighted pair group method with arithmetic mean and the neighbor-joining method clustered the accessions into three to five groups. Analysis of molecular variance indicated that 76% of the genetic variation was among individuals and 23% was between individuals. Genome-wide association analyses identified 26 SNPs associated with tan spot resistance and explained between 8.1% to 20.2% of the phenotypic variation. The SNPs were located on chromosomes 1B (1 SNP), 2B (4 SNPs), 3A (2 SNPs), 3B (2 SNPs), 4A (2 SNPs), 4B (1 SNP), 5A (2 SNPs), 5B (4 SNPs), 6A (5 SNPs), 6B (2 SNPs), and 7B (1 SNP). Four markers, one on each of chromosomes 1B, and 5A, and two on 5B, coincided with previously reported SNPs for tan spot resistance, while the remaining SNPs were either novel markers or closely related to previously reported SNPs. Eight durum wheat accessions were identified as possible novel sources of tan spot resistance that could be introgressed into elite cultivars. Conclusion: The results highlighted the significance of chromosomes 2B, 5B, and 6A as genomic regions associated with tan spot resistance.

Identification and Molecular Mapping of the Gene YrFL in Wheat Cultivar Flanders with Durable Resistance to Stripe Rust.

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most severe diseases of wheat (Triticum aestivum L.) worldwide. Identification and characterization of resistance genes is advantageous to cultivating wheat varieties with durable resistance, which is the most economic and effective strategy to control stripe rust. Flanders, a common wheat cultivar released in France in 1986, confers effective resistance to stripe rust both at the seedling and adult plant stages. To elucidate the genetic basis of resistance in Flanders, F1, F2, and F2:3 generations derived from the cross Mingxian169 × Flanders were evaluated with the most prevalent Chinese Pst race CYR33 at the seedling stage. Inheritance analysis showed that the stripe rust resistance of Flanders was controlled by a single dominant gene, temporarily designated as YrFL. Bulked segregant analysis (BSA) combined with a wheat 660K single-nucleotide polymorphism (SNP) array indicated that polymorphic SNP markers were mainly located in the 0 to 150 Mb on wheat chromosome 5A. One hundred and eleven kompetitive allele-specific PCR (KASP) and 39 simple sequence repeat (SSR) markers on chromosome 5A were used to locate the YrFL. Linkage analysis mapped YrFL with 19 KASP and three SSR markers on wheat chromosome 5AS, and the genetic distances of the closest flanking markers AX108925494 and Xbarc56 to YrFL were 0.6 and 2.0 cM, respectively. Chromosome location, resistance characterization, and molecular marker positions indicated that YrFL is likely a novel stripe rust resistance gene on wheat chromosome 5AS and could be pyramided with other resistance genes to improve resistance in wheat breeding programs.

Identification of Fusarium head blight sources of resistance and associated QTLs in historical and modern Canadian spring wheat.

Fusarium head blight (FHB) is one the most globally destructive fungal diseases in wheat and other small grains, causing a reduction in grain yield by 10-70%. The present study was conducted in a panel of historical and modern Canadian spring wheat (Triticum aestivum L.) varieties and lines to identify new sources of FHB resistance and map associated quantitative trait loci (QTLs). We evaluated 249 varieties and lines for reaction to disease incidence, severity, and visual rating index (VRI) in seven environments by artificially spraying a mixture of four Fusarium graminearum isolates. A subset of 198 them were genotyped with the Wheat 90K iSelect single nucleotide polymorphisms (SNPs) array. Genome-wide association mapping performed on the overall best linear unbiased estimators (BLUE) computed from all seven environments and the International Wheat Genome Sequencing Consortium (IWGSC) RefSeq v2.0 physical map of 26,449 polymorphic SNPs out of the 90K identified sixteen FHB resistance QTLs that individually accounted for 5.7-10.2% of the phenotypic variance. The positions of two of the FHB resistance QTLs overlapped with plant height and flowering time QTLs. Four of the QTLs (QFhb.dms-3B.1, QFhb.dms-5A.5, QFhb.dms-5A.7, and QFhb.dms-6A.4) were simultaneously associated with disease incidence, severity, and VRI, which accounted for 27.0-33.2% of the total phenotypic variance in the combined environments. Three of the QTLs (QFhb.dms-2A.2, QFhb.dms-2D.2, and QFhb.dms-5B.8) were associated with both incidence and VRI and accounted for 20.5-22.1% of the total phenotypic variance. In comparison with the VRI of the checks, we identified four highly resistant and thirty-three moderately resistant lines and varieties. The new FHB sources of resistance and the physical map of the associated QTLs would provide wheat breeders valuable information towards their efforts in developing improved varieties in western Canada.

Association Mapping of Quantitative Trait Loci for Agronomic Traits in a Winter Wheat Collection Grown in Kazakhstan

Central Asia is an important region for the growth of winter wheat, with a cultivation area of more than 15 million hectares (ha). However, the average yield is approximately 3 tons per ha, which is significantly lower than that in developed countries. Therefore, the development of new competitive high-yielding cultivars, including those based on the application of modern molecular genetics tools, is a key priority in winter wheat breeding projects. One of these tools is the result of the identification of new genes and quantitative trait loci (QTLs) for agronomic traits using diverse germplasm panels and genome-wide association studies (GWAS). In this work, a panel of winter wheat accessions was assembled using 115 accessions from Central Asia and 162 samples from other regions of the world. The GWAS, based on a two-year field evaluation of the collection in Kazakhstan’s southern and southeastern regions and 10,481 polymorphic SNP (single-nucleotide polymorphism) markers, allowed for the detection of 173 stable QTLs in nine studied agronomic traits. A survey of the published scientific literature suggests that 23 of these 173 stable QTLs have locations similar to those of previously reported QTLs, supporting the robustness of the research. In addition, 221 and 162 accessions surpassed local standards for grain yield at Kazakhstan’s southern and southeastern stations, respectively. Therefore, this study is an additional contribution to the identification of new QTLs for key agronomic traits and valuable genetic lines in winter wheat breeding projects.

Single nucleotide polymorphism analysis in Leptospermum scoparium (Myrtaceae) supports two highly differentiated endemic species in Aotearoa New Zealand and Australia

Leptospermum scoparium is the basis of a flourishing honey industry in Aotearoa New Zealand (NZ) and Australia. The genetic structure of L. scoparium across its range in NZ and Australia was previously assessed using pooled, whole genome sequencing; however, only one sampling site in Tasmania was included. Here, we used a single nucleotide polymorphism (SNP) array for genotyping samples of L. scoparium collected in natural stands around Tasmania and NZ, to determine the genetic relationship between L. scoparium individuals from the two regions. In total, 2069 high quality, polymorphic SNP markers were applied across the sample set of 504 individuals, revealing that Tasmanian L. scoparium are genetically distinct from NZ mānuka, confirming the observation from the pooled whole genome sequencing project. FST and discriminant analysis of principal components confirmed that the Tasmanian populations are well differentiated genetically from NZ populations, suggesting that they should be recognised as a separate, endemic Australian species. Within NZ, eight geographic groups are distinguished with genotypic variation exhibiting north to south landscape scale patterns with regional genetic clusters. We found support for isolation by distance, and this was reflected in the range of pairwise FST values estimated between NZ genetic clusters (0.056 to 0.356); however, each geographic genetic group exhibits geneflow and is only weakly differentiated from neighbouring clusters as evidenced by low population differentiation (low pairwise FST). These data provide little support for taxonomic revision and subdividing L. scoparium into segregate species within NZ.

Identification of salt stress-tolerant candidate genes in the BC2F2 population at the seedling stages of G. hirsutum and G. darwinii using NGS-based bulked segregant analysis.

Salinity is a major threat to the yield and productivity of cotton seedlings. In the present study, we developed a BC2F2 population of cotton plants from Gossypium darwinii (5-7) and Gossypium hirsutum (CCRI 12-4) salt-susceptible parents to identify salt-resistant candidate genes. The Illumina HiSeq™ strategy was used with bulked segregant analysis. Salt-resistant and salt-susceptible DNA bulks were pooled by using 30 plants from a BC2F2 population. Next-generation sequencing (NGS) technology was used for the sequencing of parents and both bulks. Four significant genomic regions were identified: the first genomic region was located on chromosome 18 (1.86 Mb), the second and third genomic regions were on chromosome 25 (1.06 Mb and 1.94 Mb, respectively), and the fourth was on chromosome 8 (1.41 Mb). The reads of bulk1 and bulk2 were aligned to the G. darwinii and G. hirsutum genomes, respectively, leading to the identification of 20,664,007 single-nucleotide polymorphisms (SNPs) and insertions/deletions (indels). After the screening, 6,573 polymorphic markers were obtained after filtration of the candidate regions. The SNP indices in resistant and susceptible bulks and Δ(SNP-index) values of resistant and susceptible bulks were measured. Based on the higher Δ(SNP-index) value, six effective polymorphic SNPs were selected in a different chromosome. Six effective SNPs were linked to five candidate genes in four genomic regions. Further validation of these five candidate genes was carried out using reverse transcription-quantitative polymerase chain reaction (RT-qPCR), resulting in an expression profile that showed two highly upregulated genes in the salt-tolerant species G. darwinii, i.e., Gohir.D05G367800 and Gohir.D12G239100; however, the opposite was shown in G. hirsutum, for which all genes, except one, showed partial expression. The results indicated that Gohir.D05G367800 and Gohir.D12G239100 may be salt-tolerant genes. We are confident that this study could be helpful for the cloning, transformation, and development of salt-resistant cotton varieties.

Identification of fusarium head blight resistance markers in a genome-wide association study of CIMMYT spring synthetic hexaploid derived wheat lines

Fusarium head blight (FHB), caused by Fusarium graminearum, is one of the most destructive wheat diseases worldwide. FHB infection can dramatically reduce grain yield and quality due to mycotoxins contamination. Wheat resistance to FHB is quantitatively inherited and many low-effect quantitative trait loci (QTL) have been mapped in the wheat genome. Synthetic hexaploid wheat (SHW) represents a novel source of FHB resistance derived from Aegilops tauschii and Triticum turgidum that can be transferred into common wheat (T. aestivum). In this study, a panel of 194 spring Synthetic Hexaploid Derived Wheat (SHDW) lines from the International Maize and Wheat Improvement Center (CIMMYT) was evaluated for FHB response under field conditions over three years (2017–2019). A significant phenotypic variation was found for disease incidence, severity, index, number of Fusarium Damaged Kernels (FDKs), and deoxynivalenol (DON) content. Further, 11 accessions displayed < 10 ppm DON in 2017 and 2019. Genotyping of the SHDW panel using a 90 K Single Nucleotide Polymorphism (SNP) chip array revealed 31 K polymorphic SNPs with a minor allele frequency (MAF) > 5%, which were used for a Genome-Wide Association Study (GWAS) of FHB resistance. A total of 52 significant marker-trait associations for FHB resistance were identified. These included 5 for DON content, 13 for the percentage of FDKs, 11 for the FHB index, 3 for disease incidence, and 20 for disease severity. A survey of genes associated with the markers identified 395 candidate genes that may be involved in FHB resistance. Collectively, our results strongly support the view that utilization of synthetic hexaploid wheat in wheat breeding would enhance diversity and introduce new sources of resistance against FHB into the common wheat gene pool. Further, validated SNP markers associated with FHB resistance may facilitate the screening of wheat populations for FHB resistance.

Population Genetic Analysis in Persimmons (Diospyros kaki Thunb.) Based on Genome-Wide Single-Nucleotide Polymorphisms.

This study investigated the genetic diversity and population structure of a persimmon (Diospyros kaki Thunb., 2n = 6x = 90) collection in South Korea by evaluating 9751 genome-wide single-nucleotide polymorphisms (SNPs) detected using genotyping-by-sequencing in 93 cultivars. The results of neighbor-joining clustering, principal component analysis, and STRUCTURE analysis based on SNPs indicated clear separation between cultivar groups (pollination-constant nonastringent (PCNA, 40 cultivars), pollination-constant astringent (PCA, 19), pollination-variant nonastringent (PVNA, 23), and the pollination-variant astringent type (PVA, 9)) based on the astringency types, while separation between PVA and PVNA-type cultivars was unclear. Population genetic diversity based on SNPs showed that the proportions of polymorphic SNPs within each group ranged from 99.01% (PVNA) to 94.08% (PVA), and the PVNA group exhibited the highest genetic diversity (He = 3.86 and uHe = 0.397). F (fixation index) values were low ranging from -0.024 (PVA) to 0.176 (PCA) with an average of 0.089, indicating a deficiency of heterozygosity. Analysis of molecular variance (AMOVA) and Fst among cultivar groups indicated that variation within individuals was higher than that among the groups. Pairwise Fst values among the groups ranged from 0.01566 (between PVA and PVNA) to 0.09416 (between PCA and PCNA), indicating a low level of cultivar type differentiation. These findings highlight the potential application of biallelic SNPs in population genetics studies of allopolyploids species and provide valuable insights that may have significant implications for breeding and cultivar identification in persimmon.

Genetic diversity and genome-scale population structure of wild Indian major carp, Labeo catla (Hamilton, 1822), revealed by genotyping-by-sequencing.

Labeo catla (catla) is the second most commercially important and widely cultured Indian major carp (IMC). It is indigenous to the Indo-Gangetic riverine system of India and the rivers of Bangladesh, Nepal, Myanmar, and Pakistan. Despite the availability of substantial genomic resources in this important species, detailed information on the genome-scale population structure using SNP markers is yet to be reported. In the present study, the identification of genome-wide single nucleotide polymorphisms (SNPs) and population genomics of catla was undertaken by re-sequencing six catla populations of riverine origin from distinct geographical regions. DNA isolated from 100 samples was used to perform genotyping-by-sequencing (GBS). A published catla genome with 95% genome coverage was used as the reference for mapping reads using BWA software. From a total of 472 million paired-end (150 × 2bp) raw reads generated in this study, we identified 10,485 high-quality polymorphic SNPs using the STACKS pipeline. Expected heterozygosity (He) across the populations ranged from 0.162 to 0.20, whereas observed heterozygosity (Ho) ranged between 0.053 and 0.06. The nucleotide diversity (π) was the lowest (0.168) in the Ganga population. The within-population variation was found to be higher (95.32%) than the among-population (4.68%) variation. However, genetic differentiation was observed to be low to moderate, with Fst values ranging from 0.020 to 0.084, and the highest between Brahmani and Krishna populations. Bayesian and multivariate techniques were used to further evaluate the population structure and supposed ancestry in the studied populations using the structure and discriminant analysis of principal components (DAPC), respectively. Both analyses revealed the existence of two separate genomic clusters. The maximum number of private alleles was observed in the Ganga population. The findings of this study will contribute to a deeper understanding of the population structure and genetic diversity of wild populations of catla for future research in fish population genomics.

Mapping of QTLs for morphophysiological and yield traits under water-deficit stress and well-watered conditions in maize.

Maize productivity is significantly impacted by drought; therefore, improvement of drought tolerance is a critical goal in maize breeding. To achieve this, a better understanding of the genetic basis of drought tolerance is necessary. Our study aimed to identify genomic regions associated with drought tolerance-related traits by phenotyping a mapping population of recombinant inbred lines (RILs) for two seasons under well-watered (WW) and water-deficit (WD) conditions. We also used single nucleotide polymorphism (SNP) genotyping through genotyping-by-sequencing to map these regions and attempted to identify candidate genes responsible for the observed phenotypic variation. Phenotyping of the RILs population revealed significant variability in most of the traits, with normal frequency distributions, indicating their polygenic nature. We generated a linkage map using 1,241 polymorphic SNPs distributed over 10 chromosomes (chrs), covering a total genetic distance of 5,471.55cM. We identified 27 quantitative trait loci (QTLs) associated with various morphophysiological and yield-related traits, with 13 QTLs identified under WW conditions and 12 under WD conditions. We found one common major QTL (qCW2-1) for cob weight and a minor QTL (qCH1-1) for cob height that were consistently identified under both water regimes. We also detected one major and one minor QTL for the Normalized Difference Vegetation Index (NDVI) trait under WD conditions on chr 2, bin 2.10. Furthermore, we identified one major QTL (qCH1-2) and one minor QTL (qCH1-1) on chr 1 that were located at different genomic positions to those identified in earlier studies. We found co-localized QTLs for stomatal conductance and grain yield on chr 6 (qgs6-2 and qGY6-1), while co-localized QTLs for stomatal conductance and transpiration rate were identified on chr 7 (qgs7-1 and qTR7-1). We also attempted to identify the candidate genes responsible for the observed phenotypic variation; our analysis revealed that the major candidate genes associated with QTLs detected under water deficit conditions were related to growth and development, senescence, abscisic acid (ABA) signaling, signal transduction, and transporter activity in stress tolerance. The QTL regions identified in this study may be useful in designing markers that can be utilized in marker-assisted selection breeding. In addition, the putative candidate genes can be isolated and functionally characterized so that their role in imparting drought tolerance can be more fully understood.

Diagnosis of Ovarian Carcinoma Homologous Recombination DNA Repair Deficiency From Targeted Gene Capture Oncology Assays.

Homologous recombination DNA repair deficiency (HRD) is a therapeutic biomarker for sensitivity to platinum and poly(ADP-ribose) polymerase inhibitor therapies in breast and ovarian cancers. Several molecular phenotypes and diagnostic strategies have been developed to assess HRD; however, their clinical implementation remains both technically challenging and methodologically unstandardized. We developed and validated an efficient and cost-effective strategy for HRD determination on the basis of calculation of a genome-wide loss of heterozygosity (LOH) score through targeted, hybridization capture and next-generation DNA sequencing augmented with 3,000 common, polymorphic single-nucleotide polymorphism (SNP) sites distributed genome-wide. This approach requires minimal sequence reads and can be readily integrated into targeted gene capture workflows already in use for molecular oncology. We interrogated 99 ovarian neoplasm-normal pairs using this method and compared results with patient mutational genotypes and orthologous predictors of HRD derived from whole-genome mutational signatures. LOH scores of ≥11% had >86% sensitivity for identifying tumors with HRD-causing mutations in an independent validation set (90.6% sensitivity for all specimens). We found strong agreement of our analytic approach with genome-wide mutational signature assays for determining HRD, yielding an estimated 96.7% sensitivity and 50% specificity. We observed poor concordance with mutational signatures inferred using only mutations detected by the targeted gene capture panel, suggesting inadequacy of the latter approach. LOH score did not significantly correlate with treatment outcomes. Targeted sequencing of genome-wide polymorphic SNP sites can be used to infer LOH events and subsequently diagnose HRD in ovarian tumors. The methods presented here are readily generalizable to other targeted gene oncology assays and could be adapted for HRD diagnosis in other tumor types.

Detection of 13 Hypervariable Region 1 (Hv1) SNPs using single-base extension (Sbe) primers in parallel with Sanger sequencing

The aim of this study was to determine whether single-base extension (SBE) chemistry can be applied to forensic practice of testing the target single nucleotide polymorphisms (SNPs) of the mitochondrial DNA (mtDNA) Hypervariable Region 1 (HV1). Despite itsweak discrimination power, high copy number of mtDNA per cell and its stability against degradation guarantee mtDNA testing a place in modern forensic genetics. In this research, buccal swab samples were obtained from 294 individuals from Bosnia and Herzegovina. Following DNA isolation using QIAamp® DNA Mini Kit, full sequencing of HV1 was completed using chain-termination Sanger sequencing method. SBE reactions were then performed by targeting 13 SNPs that were identified to be the most frequent in the study population. Uniplex SBE reactions for each individual SNP, as well as two multiplex reactions were prepared for both pure and mixed samples. The results showed complete agreement of the Sanger sequencing results with SBE reactions for both uniplex and multiplex reactions. The results obtained with SBE were encouraging in regard to multiplexing and processing of the mixed samples, since the allele of the minor contributor to the sample was observed in SBE electropherogram in all prepared mixtures. SBE method is limited by the fact that only target SNPs of interest will be analyzed, meaning that they must be carefully selected and curated for each population. However, typing with SBE protocol is accurate, as compared to the golden standard of Sanger sequencing, but was more time- and labor-efficient and simpler to analyze, therefore offering a suitable alternative when Sanger sequencing is not available, given that polymorphic SNPs are known for the population of interest.

Haplotype structure of MSTN, IGF1, and BMP2 genes in Tunisian goats (Capra hircus) and their association with morphometric traits.

The aim of the present study was to evaluate variability and haplotype structure of twenty-eight single nucleotide polymorphisms (SNPs) at myostatin (MSTN), insulin-like growth factor 1 (IGF1), and bone morphogenetic protein 2 (BMP2) genes. Association between the polymorphic SNPs and morphometric traits was performed on a population of 263 Tunisian goats. The SNPs analyzed were all polymorphic (except one), and the three genes had different haplotype structures. Significant association of SNPs at MSTN with head length was highlighted in the Tunisian goats. The variability at IGF1 gene was associated with body length, ear length, tail length, and chest depth. For BMP2, significant association was revealed with chest depth. Significant association was also detected between linkage disequilibrium (LD) block 2 at IGF1 with body length. These findings might play a potential role in gene-assisted programs.

Development and identification of 131 SNP markers in Sthenoteuthis pteropus (Steenstrup)

The orangeback flying squid, Sthenoteuthis pteropus, is a species of significant potential value that is widely distributed in the tropical and temperate waters of the Atlantic Ocean. There have been no reports of the population genetics and effective molecular markers for this species due to a lack of reliable information regarding its genetic structure and its many individual differences, as well as its complex and changeable life history. Therefore, the development of auxiliary molecular markers would contribute to the development, sustainable utilization, and protection of the species. In this study, 131 novel single nucleotide polymorphism (SNP) markers were developed by double digest restriction-site associated DNA sequencing (ddRAD-Seq). The observed heterozygosity (Ho) and expected heterozygosity (He) ranged from 0.00 to 0.80 and 0.18 to 0.50, respectively. The polymorphism information content (PIC) value ranged from 0.18 to 0.50. None of the marker locations significantly deviated from the Hardy–Weinberg equilibrium (p > 0.05) after a Bonferroni correction. These polymorphic SNPs will be important in the further analysis of the population heredity of S. pteropus and its scientific management.

Novel candidate loci for morpho-agronomic and seed quality traits detected by targeted genotyping-by-sequencing in common bean.

Phaseolus vulgaris L., known as common bean, is one of the most important grain legumes cultivated around the world for its immature pods and dry seeds, which are rich in protein and micronutrients. Common bean offers a cheap food and protein sources to ameliorate food shortage and malnutrition around the world. However, the genetic basis of most important traits in common bean remains unknown. This study aimed at identifying QTL and candidate gene models underlying twenty-six agronomically important traits in common bean. For this, we assembled and phenotyped a diversity panel of 200P. vulgaris genotypes in the greenhouse, comprising determinate bushy, determinate climbing and indeterminate climbing beans. The panel included dry beans and snap beans from different breeding programmes, elite lines and landraces from around the world with a major focus on accessions of African, European and South American origin. The panel was genotyped using a cost-conscious targeted genotyping-by-sequencing (GBS) platform to take advantage of highly polymorphic SNPs detected in previous studies and in diverse germplasm. The detected single nucleotide polymorphisms (SNPs) were applied in marker-trait analysis and revealed sixty-two quantitative trait loci (QTL) significantly associated with sixteen traits. Gene model identification via a similarity-based approach implicated major candidate gene models underlying the QTL associated with ten traits including, flowering, yield, seed quality, pod and seed characteristics. Our study revealed six QTL for pod shattering including three new QTL potentially useful for breeding. However, the panel was evaluated in a single greenhouse environment and the findings should be corroborated by evaluations across different field environments. Some of the detected QTL and a number of candidate gene models only elucidate the understanding of the genetic nature of these traits and provide the basis for further studies. Finally, the study showed the possibility of using a limited number of SNPs in performing marker-trait association in common bean by applying a highly scalable targeted GBS approach. This targeted GBS approach is a cost-efficient strategy for assessment of the genetic basis of complex traits and can enable geneticists and breeders to identify novel loci and targets for marker-assisted breeding more efficiently.

Identification of quantitative trait loci of agronomic traits in bread wheat using a Pamyati Azieva×Paragon mapping population harvested in three regions of Kazakhstan.

Although genome-wide association studies (GWAS) are an increasingly informative tool in the mining of new quantitative trait loci (QTLs), a classical biparental mapping approach is still a powerful, widely used method to search the unique genetic factors associated with important agronomic traits in bread wheat. In this study, a newly constructed mapping population of Pamyati Azieva (Russian Federation)×Paragon (UK), consisting of 94 recombinant inbred lines (RILs), was tested in three different regions of Kazakhstan with the purpose of QTL identification for key agronomic traits. The RILs were tested in 11 environments of two northern breeding stations (Petropavlovsk, North Kazakhstan region, and Shortandy, Aqmola region) and one southeastern station (Almalybak, Almaty region). The following eight agronomic traits were studied: heading days, seed maturation days, plant height, spike length, number of productive spikes, number of kernels per spike, thousand kernel weight, and yield per square meter. The 94 RILs of the PAxP cross were genotyped using Illumina's iSelect 20K single nucleotide polymorphism (SNP) array and resulted in the identification of 4595 polymorphic SNP markers. The application of the QTL Cartographer statistical package allowed the identification of 53 stable QTLs for the studied traits. A survey of published studies related to common wheat QTL identification suggested that 28 of those 53 QTLs were presumably novel genetic factors. The SNP markers for the identified QTLs of the analyzed agronomic traits of common wheat can be efficiently applied in ongoing breeding activities in the wheat breeding community using a marker-assisted selection approach.