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Drought stress occurred at early growth stages in wheat affecting the following growth stages. Therefore, selecting promising drought-tolerant genotypes with highly adapted traits at the seedling stage is an important task for wheat breeders and geneticists. Few research efforts were conducted on the genetic control for drought-adaptive traits at the seedling stage in wheat. In this study, a set of 146 highly diverse spring wheat core collections representing 28 different countries was evaluated under drought stress at the seedling stage. All genotypes were exposed to drought stress for 13 days by water withholding. Leaf traits including seedling length, leaf wilting, days to wilting, leaf area, and leaf rolling were scored. Moreover, root traits such as root length, maximum width, emergence angle, tip angle, and number of roots were scored. Considerable significant genetic variation was found among all genotypes tested in these experiments. The heritability estimates ranged from 0.74 (leaf witling) to 0.99 (root tip angle). A set of nine genotypes were selected and considered drought-tolerant genotypes. Among all leaf traits, shoot length had significant correlations with all root traits under drought stress. The 146 genotypes were genotyped using the Infinium Wheat 15 K single nucleotide polymorphism (SNP) array and diversity arrays technology (DArT) marker platform. The result of genotyping revealed 12,999 SNPs and 2150 DArT markers which were used to run a genome-wide association study (GWAS). The results of GWAS revealed 169 markers associated with leaf and root traits under drought stress. Out of the 169 markers, 82 were considered major quantitative trait loci (QTL). The GWAS revealed 95 candidate genes were identified with 53 genes showing evidence for drought tolerance in wheat, while the remaining candidate genes were considered novel. No shared markers were found between leaf and root traits. The results of the study provided mapping novel markers associated with new root traits at the seedling stage. Also, the selected genotypes from different countries could be employed in future wheat breeding programs not only for improving adaptive drought-tolerant traits but also for expanding genetic diversity.

Abstract Seed quality is the set of physical, genetic, and physiological characteristics, reflecting the overall germination potential. Maintaining an optimal seed quality is essential for agriculture and seed banks to preserve genetic diversity. Compared to conventional methods (e.g., germination tests), non-invasive approaches allow a more sustainable and rapid evaluation of seed quality but this is limited by high costs. The measurement of ultra-weak photon emission (UPE) and delayed fluorescence (DL), defined as biological phenomena potentially related to the physiological status of living systems, may represent a suitable approach to estimate seed quality. To test this hypothesis, seeds of five agriculturally relevant legume species (Phaseolus vulgaris L., Lathyrus sativus L., Cicer arietinum L., Pisum sativum L., and Vicia faba L.), stored at different conditions (room temperature or -18 °C) for several years, were analysed using a LIANA© prototype to collect data regarding DL and UPE occurring after UV excitation. The obtained data were integrated with germination parameters which underline species-specific behaviours in response to storage conditions. The prediction models show variable efficiency in classifying seeds based on germination which underline species-dependent links between photon emission and seed quality. Therefore, these measurements represent novel, non-invasive, and rapid approaches to evaluate seed quality.

Abstract Climate change-related droughts that recur frequently are one of the biggest obstacles to wheat (Triticum aestivum L.) productivity. Worldwide, attempts are being done to establish drought-resistant cultivars. However, progress is slow since drought tolerance is a complex trait controlled by numerous genes, and its expression is influenced by the environment. Phenotypic, biochemical physiological, and genotyping approaches are highlighted as critical research components for leveraging genetic variation in eight wheat genotypes. Treatments included eight spring wheat genotypes (IPK_040, IPK_046, IPK_050, IPK_071, IPK_105, WAS_007, WAS_024 and WAS_031), normal irrigation (NI), drought stress (D) (30% field capacity (FC)), normal irrigation with 0.5 mM SA (NSA), and drought treated with SA (DSA). The results revealed that there was a reduction in relative water content, an increase membrane leakage, and leaf chlorophyll content under drought stress. SA induced the defense responses against drought by increasing the osmolytes and the antioxidative enzymes activities. Compared to the NI group, the DSA treatment improved the water regulation, antioxidant capacity, and drought stress resistance. SA significantly reduced the deleterious effects of water stress on phenotyping more in WAS_ 024 and IPK_ 105 genotypes. The most responsive genotypes to salicylic acid were IPK_ 046 among the IPK genotypes, whereas WAS_031 genotype was amongst WAS genotypes based on the morpho-physiological traits. The findings of this study give a solid foundation for assessing drought resistance in T. aestivum and developing cultivation-specific water management methods. Graphical Abstract

IntroductionUnderstanding the dynamics of Pb tolerance across different growth stages is essential for breeding wheat varieties with comprehensive tolerance to Pb contamination throughout their life cycle. Lead (Pb) in soil affects wheat plants at all growth stages. However, the seedling stage is considered a sensitive growth stage for the tolerance of most abiotic stresses. Genetic variation in lead tolerance is poorly understood, and genetic control has still not been investigated.MaterialsIn this study, 103 highly diverse wheat genotypes were investigated under normal and lead stress conditions at the seedling stage. Different seedling growth traits were scored under each treatment. GWAS was performed via two different sets of markers: 21,750 (25K Infinium iSelect array) and 37,790 SNPs (genotyping-by-sequencing) to identify SNP markers associated with the studied seedling traits.ResultsHigh genetic variation was found among all the genotypes for all the traits. High broad-sense heritability estimates ranging from 0.25-0.93 were observed. A highly significant correlation was found among all the traits scored under normal and lead stress conditions. Low or no significant phenotypic correlations were found for lead tolerance between the seedling and adult growth stages. A set of eight genotypes were classified as lead-tolerant genotypes. The GWAS revealed a set of 222 significant markers associated with lead tolerance. Markers with pleiotropic effects were detected under Pb stress and between lead and normal conditions. Interestingly, five significant markers were found to be associated with lead tolerance at the seedling and adult growth stages.DiscussionThe results of this study provide new and novel insights into genetic control and genetic variation in lead tolerance at the early growth stage. All the genes and genotypes reported in this study will be very useful for further investigations of lead tolerance in wheat at different growth stages.

Wheat (Triticum spp.) is one of the most important cereal crops in the world. Several diseases affect wheat production and can cause 20-80% yield loss annually. Out of these diseases, stripe rust, also known as yellow rust (Puccinia striiformis f. sp. tritici), stem rust (Puccinia graminis f. sp. tritici), leaf rust (Puccinia recondita), and powdery mildew (Blumeria graminis f. sp. tritici) are the most important fungal diseases that infect the foliar part of the plant. Many efforts were made to improve wheat resistance to these diseases. Due to the continuous advancement in sequencing methods and genomic tools, genome-wide association study has become available worldwide. This analysis enabled wheat breeders to detect genomic regions controlling the resistance in specific countries. In this review, molecular markers significantly associated with the resistance of the mentioned foliar diseases in the last five years were reviewed. Common markers that control broad-spectrum resistance in different countries were identified. Furthermore, common genes controlling the resistance of more than one of these foliar diseases were identified. The importance of these genes, their functional annotation, and the potential for gene enrichment are discussed. This review will be valuable to wheat breeders in producing genotypes with broad-spectrum resistance by applying genomic selection for the target common markers and associated genes.

The genotype × environment interaction is one of the effective factors in identifying and introducing cultivars with stable grain yield in different environments. There are many statistical methods for estimating genotype × environment interaction, among which AMMI and GGE-biplot analyses provide better and more interpretable results. The objective of this study was to assess the genotype × environment interaction, as well as the adaptability and stability of 40 quinoa genotypes. The experiment was carried out in a randomized complete block design with three replications in eight environments (four locations of Iran and two years). The AMMI analysis of variance showed that the main effects of genotype and environment, as well as the interaction effect of genotype × environment were significant on grain yield. Separation of genotype × environment interaction based on the principal component method showed that the first six principal components were significant and accounted for 47.6%, 22.5%, 9%, 7%, 6% and 4.3% of the genotype × environment interaction variance, respectively. Based on the AMMI model, genotypes G16, G19, G35, G30, G39, G24, and G18 were identified as high-yielding and stable genotypes with high general adaptability. In contrast, genotypes G36, G27, G38, G9, G28, G29, G23, G34, G13, and G12 were the most unstable genotypes in the studied environments. In GGE-biplot analysis, two mega-environments were identified, and genotypes G16, G19, G25, and G17 were also identified as high-yielding and stable genotypes for these environments. Also, based on the biplot diagram of the ideal genotype, genotypes G16, G19, G17, and G35 were the nearest genotypes to the ideal genotype. In total, the results of various analyses showed that the three genotypes G16 and G19 were the superior genotypes of this experiment in terms of grain yield and stability. These genotypes can be introduced as high-yielding and stable genotypes to the climatic conditions of the studied areas.

Abstract Flag leaf (FL) dimension has been reported as a key ecophysiological aspect for boosting grain yield in wheat. A worldwide winter wheat panel consisting of 261 accessions was tested to examine the phenotypical variation and identify quantitative trait nucleotides (QTNs) with candidate genes influencing FL morphology. To this end, four FL traits were evaluated during the early milk stage under two growing seasons at the Leibniz Institute of Plant Genetics and Crop Plant Research. The results showed that all leaf traits (Flag leaf length, width, area, and length/width ratio) were significantly influenced by the environments, genotypes, and environments × genotypes interactions. Then, a genome-wide association analysis was performed using 17,093 SNPs that showed 10 novel QTNs that potentially play a role in modulating FL morphology in at least two environments. Further analysis revealed 8 high-confidence candidate genes likely involved in these traits and showing high expression values from flag leaf expansion until its senescence and also during grain development. An important QTN (wsnp_RFL_Contig2177_1500201) was associated with FL width and located inside TraesCS3B02G047300 at chromosome 3B. This gene encodes a major facilitator, sugar transporter-like, and showed the highest expression values among the candidate genes reported, suggesting their positive role in controlling flag leaf and potentially being involved in photosynthetic assimilation. Our study suggests that the detection of novel marker-trait associations and the subsequent elucidation of the genetic mechanism influencing FL morphology would be of interest for improving plant architecture, light capture, and photosynthetic efficiency during grain development.

Abstract The presence of incompatibility alleles in primary amphidiploids constitutes a reproductive barrier in newly synthesized wheat-rye hybrids. To overcome this barrier, the genome stabilization process includes large-scale chromosome rearrangements. In incompatible crosses resulting in fertile amphidiploids, the elimination of one of the incompatible alleles Eml-A1 or Eml-R1b can occur already in the somatic tissue of the wheat × rye hybrid embryo. We observed that the interaction of incompatible loci Eml-A1 of wheat and Eml-R1b of rye after overcoming embryo lethality leads to hybrid sterility in primary triticale. During subsequent seed reproductions (R1, R2 or R3) most of the chromosomes of A, B, D and R subgenomes undergo rearrangement or eliminations to increase the fertility of the amphidiploid by natural selection. Genotyping-by-sequencing (GBS) coverage analysis showed that improved fertility is associated with the elimination of entire and partial chromosomes carrying factors that either cause the disruption of plant development in hybrid plants or lead to the restoration of the euploid number of chromosomes (2n = 56) in the absence of one of the incompatible alleles. Highly fertile offspring obtained in compatible and incompatible crosses can be successfully adapted for the production of triticale pre-breeding stocks.

ABSTRACT Faba bean (Vicia faba L.) is an important pulse crop with a wide range of agroecological adaptations. The development of genomic tools and a comprehensive catalog of extant genetic diversity are crucial for developing improved faba bean cultivars. The lack of a cost‐effective genotyping platform limits the characterization of large germplasm collections, understanding of genetic diversity across populations, and implementing breeder's tools like genomic selection. Genotyping‐by‐sequencing (GBS) offers high‐resolution genotyping for both model and crop plant species, even without a reference genome sequence. The genome fragments targeted by GBS depend substantially on the restriction enzyme (RE) used for the complexity reduction step. Species with complex genomic architecture require optimization of GBS with proper RE to realize the full potential of GBS. Here, we evaluated various REs in the GBS method and identified that the combination of ApeKI/MseI proved to be the most appropriate for faba bean based on the best library quality, a high number of genomic loci spread across chromosomes, and high enrichment loci associated with the gene space. With the new optimized protocol, we constructed a genetic map using a recombinant inbred line (RIL) population and identified a QTL for seed hilum color on Chromosome 1. In addition, we also genotyped a diversity panel and performed a genome‐wide association studies (GWAS) for important agronomic traits, including plant height (PH), flowering time (FT), and number of pods per plant (PPP). We identified six SNP markers significantly associated with these traits and listed potential candidate genes. The optimized faba bean‐specific GBS procedure will facilitate access to the untapped genetic diversity for genetic research and breeding and may facilitate functional genomics.

Abstract In this study, the genetic and molecular diversity of 60 quinoa accessions was assessed using agronomically important traits related to grain yield as well as microsatellite (SSR) markers, and informative markers linked to the studied traits were identified using association study. The results showed that most of the studied traits had a relatively high diversity, but grain saponin and protein content showed the highest diversity. High diversity was also observed in all SSR markers, but KAAT023, KAAT027, KAAT036, and KCAA014 showed the highest values for most of the diversity indices and can be introduced as the informative markers to assess genetic diversity in quinoa. Population structure analysis showed that the studied population probably includes two subclusters, so that out of 60 quinoa accessions, 29 (48%) and 23 (38%) accessions were assigned to the first and second subclusters, respectively, and eight (13%) accessions were considered as the mixed genotypes. The study of the population structure using Structure software showed two possible subgroups (K = 2) in the studied population and the results of the bar plot confirmed it. Association study using the general linear model (GLM) and mixed linear model (MLM) identified the number of 35 and 32 significant marker-trait associations (MTAs) for the first year (2019) and 37 and 35 significant MTAs for the second year (2020), respectively. Among the significant MTAs identified for different traits, the highest number of significant MTAs were obtained for grain yield and 1000-grain weight with six and five MTAs, respectively.

Heavy metal pollution is a global issue that affects plant growth and human health. Copper and cadmium are two significant heavy metals that have become more concentrated in many soils. These metals are taken up by many plants, including wheat, and can cause various diseases in humans. The most effective way to mitigate the harmful effects of heavy metals is to grow tolerant wheat genotypes. In the current study, two different pot experiments were conducted to understand the genetic control of copper and cadmium tolerance in wheat seedlings. Two populations were used in this study, consisting of 92 genotypes for the copper experiment and 73 genotypes for the cadmium experiment. In both experiments, a replicated complete block design with three replications was used. Highly significant differences were found between the tested genotypes for all studied traits in both metals, except for root weight and the ratio between shoot weight and root weight under cadmium contamination. Single-marker analysis was performed for all significant traits, and a total of 265 and 381 markers were found to be significantly associated with seedling traits under copper and cadmium conditions, respectively. Of these markers, only eight were commonly associated with the tolerance to both metals. These markers were located within five different gene models that were functionally annotated to control heavy metal tolerance. Gene enrichment of the five identified genes revealed two key genes that significantly influenced eight biological processes, six molecular functions, and three Kyoto Ecyclopedia of Genes and Genomes (KEGG) pathways involved in heavy metal tolerance. The sources of the eight markers and their associated genes were identified in twelve genotypes, including one Egyptian and one Kazakhstani genotype, which showed superior responses to copper and cadmium, respectively. These genes and the genotypes carrying them are crucial for future breeding programs aimed at enhancing heavy metal tolerance in wheat.

In the context of crop breeding, plant height (PH) plays a pivotal role in determining straw and grain yield. Although extensive research has explored the genetic control of PH in wheat, there remains an opportunity for further advancements by integrating genomics with growth-related phenomics. Our study utilizes the latest genome-wide association scan (GWAS) techniques to unravel the genetic basis of temporal variation in PH across 179 Bulgarian bread wheat accessions, including landraces, tall historical, and semi-dwarf modern varieties. A GWAS was performed with phenotypic data from three growing seasons, the calculated best linear unbiased estimators, and the leveraging genotypic information from the 25K Infinium iSelect array, using three statistical methods (MLM, FarmCPU, and BLINK). Twenty-five quantitative trait loci (QTL) associated with PH were identified across fourteen chromosomes, encompassing 21 environmentally stable quantitative trait nucleotides (QTNs), and four haplotype blocks. Certain loci (17) on chromosomes 1A, 1B, 1D, 2A, 2D, 3A, 3B, 4A, 5B, 5D, and 6A remain unlinked to any known Rht (Reduced height) genes, QTL, or GWAS loci associated with PH, and represent novel regions of potential breeding significance. Notably, these loci exhibit varying effects on PH, contribute significantly to natural variance, and are expressed during seedling to reproductive stages. The haplotype block on chromosome 6A contains five QTN loci associated with reduced height and two loci promoting height. This configuration suggests a substantial impact on natural variation and holds promise for accurate marker-assisted selection. The potentially novel genomic regions harbor putative candidate gene coding for glutamine synthetase, gibberellin 2-oxidase, auxin response factor, ethylene-responsive transcription factor, and nitric oxide synthase; cell cycle-related genes, encoding cyclin, regulator of chromosome condensation (RCC1) protein, katanin p60 ATPase-containing subunit, and expansins; genes implicated in stem mechanical strength and defense mechanisms, as well as gene regulators such as transcription factors and protein kinases. These findings enrich the pool of semi-dwarfing gene resources, providing the potential to further optimize PH, improve lodging resistance, and achieve higher grain yields in bread wheat.

Abstract Unlike other growth stages of wheat, very few studies on drought tolerance have been done at the seedling stage, and this is due to the complexity and sensitivity of this stage to drought stress resulting from climate change. As a result, the drought tolerance of wheat seedlings is poorly understood and very few genes associated with drought tolerance at this stage were identified. To address this challenge, a set of 172 spring wheat genotypes representing 20 different countries was evaluated under drought stress at the seedling stage. Drought stress was applied on all tested genotypes by water withholding for 13 days. Two types of traits, namely morphological and physiological traits were scored on the leaves of all tested genotypes. Genome‐wide association study (GWAS) is one of the effective genetic analysis methods that was used to identify target single nucleotide polymorphism (SNP) markers and candidate genes for later use in marker‐assisted selection. The tested plant materials were genotyped using 25k Infinium iSelect array (25K) (herein after it will be identified as 25K) (for 172 genotypes) and genotyping‐by‐sequencing (GBS) (for 103 genotypes), respectively. The results of genotyping revealed 21,093 25K and 11,362 GBS‐SNPs, which were used to perform GWAS analysis for all scored traits. The results of GWAS revealed that 131 and 55 significant SNPs were controlling morphological and physiological traits, respectively. Moreover, a total of eight and seven SNP markers were found to be associated with more than one morphological and physiological trait under drought stress, respectively. Remarkably, 10 significant SNPs found in this study were previously reported for their association with drought tolerance in wheat. Out of the 10 validated SNP markers, four SNPs were associated with drought at the seedling stage, while the remaining six SNPs were associated with drought stress at the reproductive stage. Moreover, the results of gene enrichment revealed 18 and six pathways as highly significant biological and molecular pathways, respectively. The selection based on drought‐tolerant alleles revealed 15 genotypes with the highest number of different drought‐tolerant alleles. These genotypes can be used as candidate parents in future breeding programs to produce highly drought‐tolerant genotypes with high genetic diversity. Our findings in this study provide novel markers and useful information on the genetic basis of drought tolerance at early growth stages.

Abstract The prevalence of hypersensitivities towards wheat has increased in the last decades. Apart from celiac disease these include allergic and other inflammatory reactions summarized under the term non-celiac wheat sensitivity. One suspected trigger is the family of amylase/trypsin-inhibitors (ATIs), non-gluten proteins that are prominent wheat allergens and that activate the toll-like receptor 4 on intestinal immune cells to promote intestinal and extra-intestinal inflammation. We therefore quantified 13 ATIs in 60 German hexaploid winter wheat cultivars originating from 1891 to 2010 and harvested in three years by targeted liquid chromatography-tandem mass spectrometry combined with stable isotope dilution assay using specific marker peptides as internal standards. The total ATI content and that of the two major ATIs 0.19 and CM3 did not change from old cultivars (first registered from 1891 to 1950) to modern cultivars (1951–2010). There were also no significant changes in ATI distribution.

Net blotch (NB) and powdery mildew (PM) are major barley diseases with the potential to cause a dramatic loss in grain yield. Breeding for resistant barley genotypes in combination with identifying candidate resistant genes will accelerate the genetic improvement for resistance to NB and PM. To address this challenge, a set of 122 highly diverse barley genotypes from 34 countries were evaluated for NB and PM resistance under natural infection for in two growing seasons. Moreover, four yield traits; plant height (Ph), spike length (SL), spike weight (SW), and the number of spikelets per spike (NOS) were recorded. High genetic variation was found among genotypes in all traits scored in this study. No significant phenotypic correlation was found in the resistance between PM and NB. Immune genotypes for NB and PM were identified. A total of 21 genotypes were immune to both diseases. Of the 21 genotypes, the German genotype HOR_9570 was selected as the most promising genotype that can be used for future breeding programs. Furthermore, a genome-wide association study (GWAS) was used to identify resistant alleles to PM and NB. The results of GWAS revealed a set of 14 and 25 significant SNPs that were associated with increased resistance to PM and NB, respectively. This study provided very important genetic resources that are highly resistant to the Egyptian PM and NB pathotypes and revealed SNP markers that can be utilized to genetically improve resistance to PM and NB.

Abstract Recently, there has been a substantial increase in high‐throughput technologies that generate highly complex large datasets for use in the sciences. Plant breeding and genetics have benefited from this data explosion where many public and private institutions now implement genomic and phenomic data to predict performance thus informing germplasm selection. However, the multitude of methodologies and data generates a situation of strategic uncertainty. We set out to compare different methods of genomic and phenomic selection in the Capsicum core collection, developed through the G2P‐SOL project, producing a combination of unique and similar selected genotypes for heat tolerance. Combined, the methods tested identified a total of 33 genotypes that show tremendous promise for use as parents in heat tolerance breeding: with 13 of these being present in more than 1 selection method. Combining classical and multispectral phenotyping methods produced better selection results than either method alone. When each method was conducted without being informed by the other, similar results were obtained. Our weighted rank‐sum selection index identified 10 entries across environments that show heat tolerance, 8 of which are also selected within heat environments. This suggests that different breeding programs can reach similar results despite having different logistical constraints. Our case study within pepper germplasm using phenomic and genomic data exhibits the potential to compensate for the dearth of germplasm knowledge with high‐throughput data as well as the converse, to compensate for logistical or financial constraint to new technologies with breeder knowledge.

Genome-wide association studies (GWAS) are among the genetic tools for the mining of genomic loci associated with useful agronomic traits. The study enabled us to find new genetic markers associated with grain yield as well as quality. The sample under study consisted of spring wheat cultivars developed in different decades of the last century. A panel of 186 accessions was evaluated at VIR’s experiment station in Pushkin across a 3-year period of field trials. In total, 24 SNPs associated with six productivity characteristics were revealed. Along with detecting significant markers for each year of the field study, meta-analyses were conducted. Loci associated with useful yield-related agronomic characteristics were detected on chromosomes 4A, 5A, 6A, 6B, and 7B. In addition to previously described regions, novel loci associated with grain yield and quality were identified during the study. We presume that the utilization of contrast cultivars which originated in different breeding periods allowed us to identify new markers associated with useful agronomic characteristics.

Genetic enhancement of grain production and quality is a priority in wheat breeding projects. In this study, we assessed two key agronomic traits—grain protein content (GPC) and thousand kernel weight (TKW)—across 179 Bulgarian contemporary and historic varieties and landraces across three growing seasons. Significant phenotypic variation existed for both traits among genotypes and seasons, and no discernible difference was evident between the old and modern accessions. To understand the genetic basis of the traits, we conducted a genome-wide association study with MLM using phenotypic data from the crop seasons, best linear unbiased estimators, and genotypic data from the 25K Infinium iSelect array. As a result, we detected 16 quantitative trait nucleotides (QTNs) associated with GPC and 15 associated with TKW, all of which passed the false discovery rate threshold. Seven loci favorably influenced GPC, resulting in an increase of 1.4% to 8.1%, while four loci had a positive impact on TKW with increases ranging from 1.9% to 8.4%. While some loci confirmed previously published associations, four QTNs linked to GPC on chromosomes 2A, 7A, and 7B, as well as two QTNs related to TKW on chromosomes 1B and 6A, may represent novel associations. Annotations for proteins involved in the senescence-associated nutrient remobilization and in the following buildup of resources required for seed germination have been found for selected putative candidate genes. These include genes coding for storage proteins, cysteine proteases, cellulose-synthase, alpha-amylase, transcriptional regulators, and F-box and RWP-RK family proteins. Our findings highlight promising genomic regions for targeted breeding programs aimed at improving grain yield and protein content.

Sustainable global wheat production requires wheat varieties, that are sufficiently resistant to the main wheat diseases. The economically important fungal pathogens worldwide include powdery mildew (PM), yellow rust (YR), leaf rust (LR) and blotch causing pathogens including Septoria nodorum blotch (SNB) and Septoria tritici blotch (STB). Here, we present the evaluation of winter wheat varieties of diverse origin against the prevalent local populations of PM, YR, LR, STB and SNB under natural infection conditions through image-based phenotyping in two consecutive years (2019 and 2020). We found several varieties to be resistant against multiple diseases. Following the association mapping, we obtained a total of 206 marker trait associations for all the parameters scored which were condensed to 79 quantitative trait loci (QTLs) (eight QTLs for PM, 25 QTLs for LR, 11 QTLs for YR, 19 QTLs for SNB and eight QTLs for STB) based on the linkage disequilibrium among the molecular markers. The known genes present at these QTLs are discussed in detail. The varieties resistant to multiple diseases, identified with the QTLs and molecular markers can be considered as elite raw material for future wheat breeding.

The goal of this research is to test breeding material of different geographical origin, and material that is adapted to different agro-climatic regions. This research examines how different wheat genotypes behave in the climatic conditions of the Pannonian Plain, where environmental stress occur more and more frequently. Fifteen divergent wheat genotypes were evaluated for grain yield, heading time and yield components (spike length, grain weight per spike and plant height) across seasons 2017/2018, 2018/2019 and 2019/2020. By applying the ANOVA model, significant differences between tested traits were recorded among the analyzed genotypes. Among the genotypes there was a variability that ranged widely for all tested traits. The coefficient of variation ranged from 5.48% for plant height to 41.03% for grain weight per spike. Correlation analysis showed strong positive relationships between grain yield and grain weight per spike (r=0.87), as well as between plant height and heading time (r=0.65). Principal Component Analysis (PCA) was also applied in order to determine the differences between wheat genotypes. This yield marker study, which was carried out in the Western Balkans, could be very valuable in providing breeders with the information they need to evaluate their breeding programs.

Abstract The resistance to cereal cyst nematode (Heterodera avenae Woll.) in wheat (Triticum aestivum L.) was studied using 114 doubled haploid lines from a novel ITMI mapping population. These lines were screened for nematode infestation in a controlled environment for two years. QTL-mapping analyses were performed across two years (Y1 and Y2) as well as combining two years (CY) data. On the 114 lines that were screened, a total of 2,736 data points (genotype, batch or years, and replication combinations) were acquired. For QTL analysis, 12,093 markers (11,678 SNPs and 415 SSRs markers) were used, after filtering the genotypic data, for the QTL mapping. Composite interval mapping, using Haley-Knott regression (hk) method in R/QTL, was used for QTL analysis. In total, 19 QTLs were detected out of which 13 were novel and six were found to be colocalized or nearby to previously reported Cre genes, QTLs or MTAs for H. avenae or H. filipjevi. Nine QTLs were detected across all three groups (Y1, Y2 and CY) including a significant QTL "QCcn.ha-2D" on chromosome 2D that explains 23% of the variance. This QTL colocalized with a previously identified Cre3 locus. Novel QTL, QCcn.ha-2A, detected in the present study could be the possible unreported homeoloci to QCcn.ha-2D, QCcn.ha-2B.1 and QCcn.ha-2B.2. Six significant digenic epistatic interactions were also observed. In addition, 26 candidate genes were also identified including genes known for their involvement in PPNs (plant parasitic nematodes) resistance in different plant species. In-silico expression of putative candidate genes showed differential expression in roots during specific developmental stages. Results obtained in the present study are useful for wheat breeding to generate resistant genetic resources against H. avenae.

Quantitative trait loci (QTL) is one of the most important steps in marker-assisted selection. Few studies have validated quantitative trait loci for marker-assisted selection of yield traits under drought stress conditions in wheat. A set of 138 highly diverse wheat genotypes were tested under normal and drought stress conditions for 2 years. Plant height, heading date, spike length, grain number per spike, grain yield per spike, and 1000-kernel weight were scored. High genetic variation was found among genotypes in all traits scored under both conditions in the 2 years. The same panel was genotyped using a diversity-array technology (DArT) marker, and a genome-wide association study was performed to find alleles associated with yield traits under all conditions. A set of 191 significant DArT markers were identified in this study. The results of the genome-wide association study revealed eight common markers in wheat that were significantly associated with the same traits under both conditions in the 2 years. Out of the eight markers, seven were located on the D genome except one marker. Four validated markers were located on the 3D chromosome and found in complete linkage disequilibrium. Moreover, these four markers were significantly associated with the heading date under both conditions and the grain yield per spike under drought stress condition in the 2 years. This high-linkage disequilibrium genomic region was located within the TraesCS3D02G002400 gene model. Furthermore, of the eight validated markers, seven were previously reported to be associated with yield traits under normal and drought conditions. The results of this study provided very promising DArT markers that can be used for marker-assisted selection to genetically improve yield traits under normal and drought conditions.

Abstract Winter wheat growing areas in the Northern hemisphere are regularly exposed to heavy frost. Due to the negative impact on yield, the identification of genetic factors controlling frost tolerance (FroT) and development of tools for breeding is of prime importance. Here, we detected QTL associated with FroT by genome wide association studies (GWAS) using a diverse panel of 276 winter wheat genotypes that was phenotyped at five locations in Germany and Russia in three years. The panel was genotyped using the 90 K iSelect array and SNPs in FroT candidate genes. In total, 17,566 SNPs were used for GWAS resulting in the identification of 53 markers significantly associated (LOD ≥ 4) to FroT, corresponding to 23 QTL regions located on 11 chromosomes (1A, 1B, 2A, 2B, 2D, 3A, 3D, 4A, 5A, 5B and 7D). The strongest QTL effect confirmed the importance of chromosome 5A for FroT. In addition, to our best knowledge, eight FroT QTLs were discovered for the first time in this study comprising one QTL on chromosomes 3A, 3D, 4A, 7D and two on chromosomes 1B and 2D. Identification of novel FroT candidate genes will help to better understand the FroT mechanism in wheat and to develop more effective combating strategies.

Abstract Introgressions from crop wild relatives (CWRs) have been used to introduce beneficial traits into cultivated plants. Introgressions have traditionally been detected using cytological methods. Recently, single nucleotide polymorphism (SNP)-based methods have been proposed to detect introgressions in crosses for which both parents are known. However, for unknown material, no method was available to detect introgressions and predict the putative donor species. Here, we present a method to detect introgressions and the putative donor species. We demonstrate the utility of this method using 10 publicly available wheat genome sequences and identify nine major introgressions. We show that the method can distinguish different introgressions at the same locus. We trace introgressions to early wheat cultivars and show that natural introgressions were utilised in early breeding history and still influence elite lines today. Finally, we provide evidence that these introgressions harbour resistance genes.

Drought is a major constraint in wheat (Triticum aestivum L.) grain yield. The present work aimed to identify quantitative trait nucleotides (QTNs)/ candidate genes influencing drought tolerance-related traits at the seedling stage in 261 accessions of a diverse winter wheat panel. Seeds from three consecutive years were exposed to polyethylene glycol 12% (PEG-6000) and a control treatment (distilled water). The Farm-CPU method was used for the association analysis with 17,093 polymorphic SNPs. PEG treatment reduced shoot length (SL) (-36.3%) and root length (RL) (-11.3%) compared with control treatments, while the coleoptile length (CL) was increased by 11% under drought conditions, suggesting that it might be considered as an indicator of stress-tolerance. Interestingly, we revealed 70 stable QTN across 17 chromosomes. Eight QTNs related to more than one trait were detected on chromosomes 1B, 2A (2), 2B, 2D, 4B, 7A, and 7B and located nearby or inside candidate genes within the linkage disequilibrium (LD) interval. For instance, the QTN on chromosome 2D is located inside the gene TraesCS2D02G133900 that controls the variation of CL_S and SL_C. The allelic variation at the candidate genes showed significant influence on the associated traits, demonstrating their role in controlling the natural variation of multi-traits of drought stress tolerance. The gene expression of these candidate genes under different stress conditions validates their biological role in stress tolerance. Our findings offer insight into understanding the genetic factors and diverse mechanisms in response to water shortage conditions that are important for wheat improvement and adaptation at early developmental stages.

Societal Impact Statement To feed the world's ever‐increasing population, new genetic approaches are required. Increasing the number of living spikelets is one promising way to improve grain yield. This, in turn, increases the number of spikelets per plant, thereby increasing the total yield. We present the first evidence for genetic control of alive spikelets in barley. Discovering natural variation as well as genomic regions associated with these traits will serve as a benchmark in future breeding for improving grain yield. Summary The primary goal of most breeding programmes is to increase grain yield. However, one of the many methods for raising yield that is yet to be fully investigated is increasing the number of spikelets by minimising spikelet abortion. Spikelet abortion dramatically increases during the late reproductive phase, but the molecular and genetic mechanisms remain unknown. Here, we employed a phenotyping approach in which developed and undeveloped spikelets were detected and counted during spike development and their maximum yield potential (MYP) was investigated. We studied 20 agronomic and spikelet‐related traits using a set of 184 diverse spring barley accessions under field conditions. By employing a set of >125K SNPs, GWAS was conducted. Our analysis revealed 26 genetic clusters associated with MYP and the number of developed and undeveloped spikelets. Most of the significant associated genomic regions were co‐located near the candidate genes of phytohormones such as ABA, auxin, and cytokinin suggesting the importance of phytohormones in keeping spikelets alive, their development, and MYP. Our findings point to a potential link between jasmonic acid and the MYP, development and abortion of spikelets. We further provide genetic evidence that sugar‐related genes and sucrose have the potential to regulate MYP, spikelet development and spikelet survival. Our findings can be used for marker‐assisted breeding and as a resource for future molecular and genetic validation. Collectively, we propose a new genetic network linking spikelet‐related traits to grain yield determinants.

High seed quality is key to agricultural production, which is increasingly affected by climate change. We studied the effects of drought and elevated temperature during seed production on key seed quality traits of two genotypes of malting barley (Hordeum sativum L.). Plants of a “Hana-type” landrace (B1) were taller, flowered earlier and produced heavier, larger and more vigorous seeds that resisted ageing longer compared to a semi-dwarf breeding line (B2). Accordingly, a NAC domain-containing transcription factor (TF) associated with rapid response to environmental stimuli, and the TF ABI5, a key regulator of seed dormancy and vigour, were more abundant in B1 seeds. Drought significantly reduced seed yield in both genotypes, and elevated temperature reduced seed size. Genotype B2 showed partial thermodormancy that was alleviated by drought and elevated temperature. Metabolite profiling revealed clear differences between the embryos of B1 and B2. Drought, but not elevated temperature, affected the metabolism of amino acids, organic acids, osmolytes and nitrogen assimilation, in the seeds of both genotypes. Our study may support future breeding efforts to produce new lodging and drought resistant malting barleys without trade-offs that can occur in semi-dwarf varieties such as lower stress resistance and higher dormancy.

Most of the freshwaters worldwide are used for agriculture. Freshwater sources are expected to decline and will not suffice to support the food production needed for the growing population. Therefore, growing crops with seawater might constitute a solution. However, very little work has been done on the effect of seawater stress on wheat, an important cereal crop. The present study aimed to determine whether particular wheat genotypes provided better resistance to seawater stress. A set of 80 highly diverse spring wheat genotypes collected from different countries in Europe, Asia, Africa, North and South America was exposed to 50% seawater stress at the early growth stage. Four seeding shoot and root traits were scored for all genotypes. High genetic variations were found among all genotypes for the epicotyl length (EL), hypocotyl length (HL), number of radicles (NOR), and fresh weight (FW). Eight genotypes with high-performance scores of seedling traits were selected. The correlation analyses revealed highly significant correlations among all traits scored in this study. The strongest correlation was found between the NOR and the other seeding traits. Thus, the NOR might be an important adaptive trait for seawater tolerance. The genetic diversity among all genotypes was investigated based on genetic distance. A wide range of genetic distances among all genotypes was found. There was also a great genetic distance among the eight selected genotypes. In particular, the genetic distance between ATRI 5310 (France) and the other seven genotypes was the greatest. Such high genetic diversity might be utilized to select highly divergent genotypes for crossing in a future breeding program. The present study provides very useful information on the presence of different genetic resources in wheat for seawater tolerance.

Drought is one of the complex abiotic stresses that affect the growth and production of wheat in arid and semiarid countries. In this study, a set of 172 diverse spring wheat genotypes from 20 different countries were assessed under drought stress at the seedling stage. Besides seedling length, two types of traits were recorded, namely: tolerance traits (days to wilting, leaf wilting, and the sum of leaf wilting), and recovery traits (days to regrowth, regrowth biomass, and drought survival rate). In addition, tolerance index, recovery index, and drought tolerance index (DTI) were estimated to select the most drought tolerant genotypes. Moreover, leaf protein content (P), amino acid (AM), proline content (PRO), glucose (G), fructose (F), and total soluble carbohydrates (TSC) were measured under control and drought conditions to study the changes in each physiological trait due to drought stress. All genotypes showed a high significant genetic variation in all the physio-morphological traits scored under drought stress. High phenotypic and genotypic correlations were found among all seedling morphological traits. Among the studied indices, the drought tolerance index (DTI) had the highest phenotypic and genotypic correlations with all tolerance and recovery traits. The broad-sense heritability (H2) estimates were high for morphological traits (83.85–92.27), while the physiological traits ranged from 96.41 to 98.68 under the control conditions and from 97.13 to 99.99 under drought stress. The averages of the physiological traits (proteins, amino acids, proline, glucose, fructose, and total soluble carbohydrates) denoted under drought stress were higher than those recorded under well-watered conditions except for proteins. In this regard, amino acids, glucose, and total soluble carbohydrates had a significant correlation with all morphological traits. The selection for drought tolerance revealed 10 tolerant genotypes from different countries (8 genotypes from Egypt, one from Morocco, and one from the United States). These selected genotypes were screened for the presence of nine specific TaDREB1 alleles. Six primers were polymorphic among the selected genotypes. Genetic diversity among the selected genotypes was investigated using 21,450 SNP markers. The results of the study shed light on the different mechanisms for drought tolerance that wheat plants use to tolerate and survive under drought stress. The genetic analysis performed in this study suggested the most suitable genotypes for selective breeding at the seedling stage under water deficit.

Abstract Background The future productivity of wheat (T. aestivum L.) as the most grown crop worldwide is of utmost importance for global food security. Thousand kernel weight (TKW) in wheat is closely associated with grain architecture-related traits, e.g. kernel length (KL), kernel width (KW), kernel area (KA), kernel diameter ratio (KDR), and factor form density (FFD). Discovering the genetic architecture of natural variation in these traits, identifying QTL and candidate genes are the main aims of this study. Therefore, grain architecture-related traits in 261 worldwide winter accessions over three field-year experiments were evaluated. Results Genome-wide association analysis using 90K SNP array in FarmCPU model revealed several interesting genomic regions including 17 significant SNPs passing false discovery rate threshold and strongly associated with the studied traits. Four of associated SNPs were physically located inside candidate genes within LD interval e.g. BobWhite_c5872_589 (602,710,399 bp) found to be inside TraesCS6A01G383800 (602,699,767–602,711,726 bp). Further analysis reveals the four novel candidate genes potentially involved in more than one grain architecture-related traits with a pleiotropic effects e.g. TraesCS6A01G383800 gene on 6A encoding oxidoreductase activity was associated with TKW and KA. The allelic variation at the associated SNPs showed significant differences betweeen the accessions carying the wild and mutated alleles e.g. accessions carying C allele of BobWhite_c5872_589, TraesCS6A01G383800 had significantly higher TKW than the accessions carying T allele. Interestingly, these genes were highly expressed in the grain-tissues, demonstrating their pivotal role in controlling the grain architecture. Conclusions These results are valuable for identifying regions associated with kernel weight and dimensions and potentially help breeders in improving kernel weight and architecture-related traits in order to increase wheat yield potential and end-use quality.

Wheat stripe rust (WSR) caused by Puccinia striiformis F. tritici Erikss. (Pst) is one of the serious diseases that affect wheat planting areas around the world. Many efforts have been made to control such a serious disease including using fungicides and breeding highly resistant genotypes. However, due to Pst’s ability to produce new races that overcome these fungicides and break the resistance in the highly resistant genotypes, looking for other effective ways to restrict this disease is urgently required. One of the highly effective ways of controlling crop diseases is using biological control. In this study, the efficiency of three different Trichoderma species (Trichoderma asperellum T34, Trichoderma harzianum (TH), and Trichoderma verdinium (TV)) was tested in a set of 34 wheat genotypes at the seedling stage. The evaluation was conducted in two experiments with two different temperature regimes. In each experiment, four treatments were applied, namely, control, T34, TV, and TH. High genetic variation was found among all genotypes in each experiment and under each Trichoderma treatment. Notably, the symptoms of WSR were affected by temperature under all treatments except T34, which had a stable performance in the two experiments. The 34 studied genotypes were highly diverse, related to ten different countries, and consisted of durum and bread wheat. Out of the three studied Trichoderma species, T34 was able to improve WSR resistance in all the studied genotypes suggesting its effectiveness in inducing the resistance and producing a priming response in different wheat genetic backgrounds. The results of this study provided very useful information on the effectiveness of Trichoderma spp. in controlling WSR.

Natalia Tikhenko et al. investigate the genetic contribution of the wheat chromosome 1D to its development and evolution. They find a novel locus, DEFECTIVE ENDOSPERM-D1, on the long arm of 1D that is required for normal endosperm development as its absence leads to non-viable grains and altered grain shape.

Abstract Background Quantitative PCR (qPCR) is one of the most common and accurate methods of gene expression analysis. However, the biggest challenge for this kind of examinations is normalization of the results, which requires the application of dependable internal controls. The selection of appropriate reference genes (RGs) is one of the most crucial points in qPCR data analysis and for correct assessment of gene expression. Because of the fact that many reports indicate that the expression profiles of typically used RGs can be unstable in certain experimental conditions, species or tissues, reference genes with stable expression levels should be selected individually for each experiment. In this study, we analysed a set of ten candidate RGs for wheat seedlings under short-term drought stress. Our tests included five ‘traditional’ RGs (GAPDH, ACT, UBI, TUB, and TEF1) and five novel genes developed by the RefGenes tool from the Genevestigator database. Results Expression stability was assessed using five different algorithms: geNorm, NormFinder, BestKeeper, RefFinder and the delta Ct method. In the final ranking, we identified three genes: CJ705892, ACT, and UBI, as the best candidates for housekeeping genes. However, our data indicated a slight variation between the different algorithms that were used. We revealed that the novel gene CJ705892, obtained by means of in silico analysis, showed the most stable expression in the experimental tissue and condition. Conclusions Our results support the statement, that novel genes selected for certain experimental conditions have a more stable level of expression in comparison to routinely applied RGs, like genes encoding actin, tubulin or GAPDH. Selected CJ705892 gene can be used as a housekeeping gene in the expression analysis in wheat seedlings under short-term drought. The results of our study will be useful for subsequent analyses of gene expression in wheat tissues subjected to drought.

Wheat leaf rust (LR) causes significant yield losses worldwide. In Egypt, resistant cultivars began to lose their efficiency in leaf rust resistance. Therefore, a diverse spring wheat panel was evaluated at the seedling stage to identify new sources of broad-spectrum seedling resistance against the Egyptian Puccinia triticina (Pt) races. In three different experiments, seedling evaluation was done using Pt spores collected from different fields and growing seasons. Highly significant differences were found among experiments confirming the presence of different races population in each experiment. Highly significant differences were found among the tested genotypes confirming the ability to select superior genotypes. Genome-wide association study (GWAS) was conducted for each experiment and a set of 87 markers located within 48 gene models were identified. The identified gene models were associated with disease resistance in wheat. Five gene models were identified to resist all Pt races in at least two experiments and could be identified as stable genes under Egyptian conditions. Ten genotypes from five different countries were stable against all the tested Pt races but showed different degrees of resistance.

Wheat stripe rust (Puccinia striiformis Westend f. sp. tritici; Pst) is a severe disease that affects wheat-growing areas worldwide. In the last few years, Pst head infection (PstHI) was recorded to cause severe yield damage in Egypt. Little information is known about PstHI. In the current study, a set of 34 Egyptian cultivars were evaluated for their Pst foliar infection (PstFI) and PstHI in two growing seasons (2021 and 2022) in Egypt. High genotypic variations in PstFI and PstHI were found. A very weak correlation was found between the resistance to PstFI and PstHI in both growing seasons. Therefore, the resistance to each symptom might be controlled by different genetic systems. The effect of PstFI and PstHI on kernel traits was investigated. PstHI was found to significantly reduce kernel width and weight in heavily diseased years. Therefore, a significant reduction in wheat yield and quality is expected in case of severe PstHI. Three Egyptian genotypes were resistant to PstFI and PstHI in both growing seasons. These genotypes could help in improving the resistance to both symptoms. However, due to the low genetic distance among them, looking for additional sources of resistance is recommended. In 2021, higher head infection was found compared with 2022. Highly significant differences in maximum temperature, minimum temperature, maximum dew point, and average dew point were found between the two growing seasons. Out of these four factors, dew point and ambient temperature were found to be possible reasons for increasing PstHI. Furthermore, 51 isolines carrying different resistance genes were included in the evaluation. At least ten stripe rust resistance genes were found to be effective against PstFI and PstHI. These ten genes should be included in future breeding programs to improve Pst resistance and understand the effect of climate change on Pst.

Climate change is an undeniable threat to sustainable wheat production in the future as an increased temperature will significantly increase grain loss due to the increased number of generations per season of multivoltine species that are detrimental to plants. Among insects, orange wheat blossom midge (OWBM), yellow wheat blossom midge (YWBM), saddle gall midge (SGM), thrips, and frit fly (FF) are important wheat pests in the European environments, which can be managed by the development of resistant cultivars. This involves the identification, confirmation, and incorporation of insect resistance sources into new high-yielding cultivars. We used two diverse and unrelated wheat [winter wheat (WW) and spring wheat (SW)] panels to associate single-nucleotide polymorphism (SNP) markers with the mentioned pests using the tools of association mapping. All in all, a total of 645 and 123 significant associations were detected in WW and SW, respectively, which were confined to 246 quantitative trait loci. Many candidate genes were identified using the BLAST analysis of the sequences of associated SNPs. Some of them are involved in controlling the physical structures of plants such as stomatal immunity and closure, cuticular wax in leaf blade, whereas others are involved in the production of certain enzymes in response to biotic and abiotic stresses. To our knowledge, this is the first detailed investigation that deals with YWBM, SGM, thrips, and FF resistance genetics using the natural variation in wheat. The reported germplasm is also readily available to breeders across the world that can make rational decisions to breed for the pest resilience of their interest by including the resistant genotypes being reported.

Seed longevity is the most important trait in the genebank management system. No seed can remain infinitely viable. There are 1241 accessions of Capsicum annuum L. available at the German Federal ex situ genebank at IPK Gatersleben. C. annuum (Capsicum) is the most economically important species of the genus Capsicum. So far, there is no report that has addressed the genetic basis of seed longevity in Capsicum. Here, we convened a total of 1152 Capsicum accessions that were deposited in Gatersleben over forty years (from 1976 to 2017) and assessed their longevity by analyzing the standard germination percentage after 5–40 years of storage at −15/−18 °C. These data were used to determine the genetic causes of seed longevity, along with 23,462 single nucleotide polymorphism (SNP) markers covering all of the 12 Capsicum chromosomes. Using the association-mapping approach, we identified a total of 224 marker trait associations (MTAs) (34, 25, 31, 35, 39, 7, 21 and 32 MTAs after 5-, 10-, 15-, 20-, 25-, 30-, 35- and 40-year storage intervals) on all the Capsicum chromosomes. Several candidate genes were identified using the blast analysis of SNPs, and these candidate genes are discussed.

Grain protein content (GPC) is a key aspect of grain quality, a major determinant of the flour functional properties and grain nutritional value of bread wheat. Exploiting diverse germplasms to identify genes for improving crop performance and grain nutritional quality is needed to enhance food security. Here, we evaluated GPC in a panel of 255 Triticum aestivum L. accessions from 27 countries. GPC determined in seeds from three consecutive crop seasons varied from 8.6 to 16.4% (11.3% on average). Significant natural phenotypic variation in GPC among genotypes and seasons was detected. The population was evaluated for the presence of the trait-linked single nucleotide polymorphism (SNP) markers via a genome-wide association study (GWAS). GWAS analysis conducted with calculated best linear unbiased estimates (BLUEs) of phenotypic data and 90 K SNP array using the fixed and random model circulating probability unification (FarmCPU) model identified seven significant genomic regions harboring GPC-associated markers on chromosomes 1D, 3A, 3B, 3D, 4B and 5A, of which those on 3A and 3B shared associated SNPs with at least one crop season. The verified SNP–GPC associations provide new promising genomic signals on 3A (SNPs: Excalibur_c13709_2568 and wsnp_Ku_c7811_13387117) and 3B (SNP: BS00062734_51) underlying protein improvement in wheat. Based on the linkage disequilibrium for significant SNPs, the most relevant candidate genes within a 4 Mbp-window included genes encoding a subtilisin-like serine protease; amino acid transporters; transcription factors; proteins with post-translational regulatory functions; metabolic proteins involved in the starch, cellulose and fatty acid biosynthesis; protective and structural proteins, and proteins associated with metal ions transport or homeostasis. The availability of molecular markers within or adjacent to the sequences of the detected candidate genes might assist a breeding strategy based on functional markers to improve genetic gains for GPC and nutritional quality in wheat.

Understanding the genetic complexity of traits is an important objective of small grain temperate cereals yield and adaptation improvements. Bi-parental quantitative trait loci (QTL) linkage mapping is a powerful method to identify genetic regions that co-segregate in the trait of interest within the research population. However, recently, association or linkage disequilibrium (LD) mapping using a genome-wide association study (GWAS) became an approach for unraveling the molecular genetic basis underlying the natural phenotypic variation. Many causative allele(s)/loci have been identified using the power of this approach which had not been detected in QTL mapping populations. In barley (Hordeum vulgare L.), GWAS has been successfully applied to define the causative allele(s)/loci which can be used in the breeding crop for adaptation and yield improvement. This promising approach represents a tremendous step forward in genetic analysis and undoubtedly proved it is a valuable tool in the identification of candidate genes. In this review, we describe the recently used approach for genetic analyses (linkage mapping or association mapping), and then provide the basic genetic and statistical concepts of GWAS, and subsequently highlight the genetic discoveries using GWAS. The review explained how the candidate gene(s) can be detected using state-of-art bioinformatic tools. Keywords: Association mapping, Hordeum vulgare L, Gene identification, QTL mapping, Barley breeding, GWAS

Abstract Background Salinity is a significant environmental stress factor limiting crops productivity. Barley (Hordeum vulgare L.) has a natural tolerance to salinity stress, making it an interesting study object in stress biology research. In the present study, for the first time the effect of salinity stress on barley inflorescence developmental stages was investigated. Five spring barley genotypes irrigated with saline water (12.5 ds/m NaCl) were compared to controls treated with normal tap water. We measured abscisic acid (ABA) concentrations in the apical, central and basal sections of the immature inflorescence at green anther (GA) stage. The role of ABA in spikelet primordia development, atrophy and abortion and final yield was evaluated. Results A time course experiment starting from double ridge until green anther (GA) stages revealed that salinity reduced the length of spike developmental stages in all genotypes causing shortened of the plant life cycle. The shortened plant life cycle negatively affected plant height and number of tillers/plant. Salinity also affected spikelet primordia development. In both control and salinity treated plants apical spikelet abortion started in late awn primordium (AP) stage. However, under salinity treatment, significantly more spikelets were aborted, thus directly affecting plant yield potential. ABA, which plays a role in the spikelet/floret abortion process, was markedly elevated in the base and apex of salt treated spikes correlating with an increased spikelet abortion in these regions. Conclusions Overall, salinity treatment reduced all plant and yield-related parameters investigated and turned some of the correlations among them from positive to negative or vice versa. Investigations of ABA role in floral development and phase duration of barley spike showed that, ABA regulates the spikelet/floret abortion process affecting the yield potential under salinity and control conditions.

The critical node (CN, 85% germination) of seed viability is an important threshold for seed regeneration decisions after long-term conservation. Dependent on the germplasm, the storage period until CN is reached varies and information on the divergence of the proteomic profiles is limited. Therefore, the study aims to identify key proteins and mechanisms relevant for a long plateau phase and a late CN during artificial seed aging of wheat. Seeds of the storage-tolerant genotype (ST) TRI 23248, and the storage-sensitive genotype (SS) TRI 10230 were exposed to artificial ageing (AA) and extracted embryos of imbibed seeds were analyzed using an iTRAQ-based proteomic technique. ST and SS required AA for 24 and 18 days to reach the CN, respectively. Fifty-seven and 165 differentially abundant proteins (DAPs) were observed in the control and aged groups, respectively. Interestingly, a higher activity in metabolic processes, protein synthesis, transcription, cell growth/division, and signal transduction were already found in imbibed embryos of control ST seeds. After AA, 132 and 64 DAPs were accumulated in imbibed embryos of both aged ST and SS seeds, respectively, which were mainly associated with cell defense, rescue, and metabolism. Moreover, 78 DAPs of ST appeared before CN and were mainly enriched in biological pathways related to the maintenance of redox and carbon homeostasis and they presented a stronger protein translation ability. In contrast, in SS, only 3 DAPs appeared before CN and were enriched only in the structural constituents of the cytoskeleton. In conclusion, a longer span of plateau phase might be obtained in seeds when proteins indicate an intense stress response before CN and include the effective maintenance of cellular homeostasis, and avoidance of excess accumulation of cytotoxic compounds. Although key proteins, inherent factors and the precise regulatory mechanisms need to be further investigated, the found proteins may also have functional potential roles during long-term seed conservation.

The size, shape, and color of wheat seeds are important traits that are associated with yield and flour quality (size, shape), nutritional value, and pre-harvest sprouting (coat color). These traits are under multigenic control, and to dissect their molecular and genetic basis, quantitative trait loci (QTL) analysis is used. We evaluated 114 recombinant inbred lines (RILs) in a bi-parental RIL mapping population (the International Triticeae Mapping Initiative, ITMI/MP) grown in 2014 season. We used digital image analysis for seed phenotyping and obtained data for seven traits describing seed size and shape and 48 traits of seed coat color. We identified 212 additive and 34 pairs of epistatic QTLs on all the chromosomes of wheat genome except chromosomes 1A and 5D. Many QTLs were overlapping. We demonstrated that the overlap between QTL regions was low for seed size/shape traits and high for coat color traits. Using the literature and KEGG data, we identified sets of genes in Arabidopsis and rice from the networks controlling seed size and color. Further, we identified 29 and 14 candidate genes for seed size-related loci and for loci associated with seed coat color, respectively.

Wheat (Triticum aestivum L.) is one of the most important crops in the world. Four spike-related traits, namely, spike weight (SW), spike length (SL), the total number of spikelets per spike (TSNS), total kernels per spike (TKNS), and thousand-kernel weight (TKW), were evaluated in 270 F3:6 Nebraska winter wheat lines in two environments (Lincoln and North Platte, NE, USA). All genotypes in both locations exhibited high genetic variation for all yield traits. High positive correlations were observed among all yield-related traits in each location separately. No or low correlation in yield-related traits was observed between the two environments. The broad-sense heritability estimates were 72.6, 72.3, 71.2, 72.3, and 56.1% for SW, SL, TSNS, TKNS, and TKW, respectively. A genome-wide association study (GWAS) was used to identify SNPs associated with yield traits. In the Lincoln environment, 44 markers were found to be significantly associated with spike-related traits (SW, SL, TSNS, TKNS, and TKW), while 41 were detected in North Platte. Due to the strong significant genotype x environment, no common SNP markers were found between the two locations. Gene annotation of the significant markers revealed candidate genes encoded for important proteins that are associated directly or indirectly with yield traits. Such high genetic variation among genotypes is very useful for selection to improve yield traits in each location separately.

Abstract Background Understanding the genetic basis of frost tolerance (FT) in wheat (Triticum aestivum L.) is essential for preventing yield losses caused by frost due to cellular damage, dehydration and reduced metabolism. FT is a complex trait regulated by a number of genes and several gene families. Availability of the wheat genomic sequence opens new opportunities for exploring candidate genes diversity for FT. Therefore, the objectives of this study were to identity SNPs and insertion-deletion (indels) in genes known to be involved in frost tolerance and to perform association genetics analysis of respective SNPs and indels on FT. Results Here we report on the sequence analysis of 19 candidate genes for FT in wheat assembled using the Chinese Spring IWGSC RefSeq v1.0. Out of these, the tandem duplicated C-repeat binding factors (CBF), i.e. CBF-A3, CBF-A5, CBF-A10, CBF-A13, CBF-A14, CBF-A15, CBF-A18, the vernalisation response gene VRN-A1, VRN-B3, the photoperiod response genes PPD-B1 and PPD-D1 revealed association to FT in 235 wheat cultivars. Within six genes (CBF-A3, CBF-A15, VRN-A1, VRN-B3, PPD-B1 and PPD-D1) amino acid (AA) substitutions in important protein domains were identified. The amino acid substitution effect in VRN-A1 on FT was confirmed and new AA substitutions in CBF-A3, CBF-A15, VRN-B3, PPD-B1 and PPD-D1 located at highly conserved sites were detected. Since these results rely on phenotypic data obtained at five locations in 2 years, detection of significant associations of FT to AA changes in CBF-A3, CBF-A15, VRN-A1, VRN-B3, PPD-B1 and PPD-D1 may be exploited in marker assisted breeding for frost tolerance in winter wheat. Conclusions A set of 65 primer pairs for the genes mentioned above from a previous study was BLASTed against the IWGSC RefSeq resulting in the identification of 39 primer combinations covering the full length of 19 genes. This work demonstrates the usefulness of the IWGSC RefSeq in specific primer development for highly conserved gene families in hexaploid wheat and, that a candidate gene association genetics approach based on the sequence data is an efficient tool to identify new alleles of genes important for the response to abiotic stress in wheat.

Seed longevity is the most important trait related to the management of gene banks because it governs the regeneration cycle of seeds. Thus, seed longevity is a quantitative trait. Prior to the discovery of molecular markers, classical genetic studies have been performed to identify the genetic determinants of this trait. Post-2000 saw the use of DNA-based molecular markers and modern biotechnological tools, including RNA sequence (RNA-seq) analysis, to understand the genetic factors determining seed longevity. This review summarizes the most important and relevant genetic studies performed in Arabidopsis (24 reports), rice (25 reports), barley (4 reports), wheat (9 reports), maize (8 reports), soybean (10 reports), tobacco (2 reports), lettuce (1 report) and tomato (3 reports), in chronological order, after discussing some classical studies. The major genes identified and their probable roles, where available, are debated in each case. We conclude by providing information about many different collections of various crops available worldwide for advanced research on seed longevity. Finally, the use of new emerging technologies, including RNA-seq, in seed longevity research is emphasized by providing relevant examples.

Wheat stripe rust (caused by Puccinia striiformis f. sp. tritici) is a major disease that damages wheat plants and affects wheat yield all over the world. In recent years, stripe rust became a major problem that affects wheat yield in Egypt. New races appeared and caused breakdowns in the resistant genotypes. To improve resistance in the Egyptian genotypes, new sources of resistance are urgently needed. In the recent research, a set of 95 wheat genotypes collected from 19 countries, including Egypt, were evaluated for their resistance against the Egyptian race(s) of stripe rust under field conditions in the two growing seasons 2018/2019 and 2019/2020. A high genetic variation was found among the tested genotypes. Single marker analysis was conducted using a subset of 71 genotypes and 424 diversity array technology (DArT) markers, well distributed across the genome. Out of the tested markers, 13 stable markers were identified that were significantly associated with resistance in both years (p-value ≤ 0.05). By using the sequence of the DArT markers, the chromosomal position of the significant DArT markers was detected, and nearby gene models were identified. Two markers on chromosomes 5A and 5B were found to be located within gene models functionally annotated with disease resistance in plants. These two markers could be used in marker-assisted selection for stripe rust resistance under Egyptian conditions. Two German genotypes were carrying the targeted allele of all the significant DArT markers associated with stripe rust resistance and could be used to improve resistance under Egyptian conditions.

MADS-box transcription factors are crucial regulators of inflorescence and flower development in plants. Therefore, the recent interest in this family has received much attention in plant breeding programs due to their impact on plant development and inflorescence architecture. The aim of this study was to investigate the role of HvMADS-box genes in lateral spikelet development in barley (Hordeum vulgare L.). A set of 30 spike-contrasting barley lines were phenotypically and genotypically investigated under controlled conditions. We detected clear variations in the spike and spikelet development during the developmental stages among the tested lines. The lateral florets in the deficiens and semi-deficiens lines were more reduced than in two-rowed cultivars except cv. Kristina. Interestingly, cv. Kristina, int-h.43 and int-i.39 exhibited the same behavior as def.5, def.6, semi-def.1, semi-def.8 regarding development and showed reduced lateral florets size. In HOR1555, HOR7191 and HOR7041, the lateral florets continued their development, eventually setting seeds. In contrast, lateral florets in two-rowed barley stopped differentiating after the awn primordia stage giving rise to lateral floret sterility. At harvest, the lines tested showed large variation for all central and lateral spikelet-related traits. Phylogenetic analysis showed that more than half of the 108 MADS-box genes identified are highly conserved and are expressed in different barley tissues. Re-sequence analysis of a subset of these genes showed clear polymorphism in either SNPs or in/del. Variation in HvMADS56 correlated with altered lateral spikelet morphology. This suggests that HvMADS56 plays an important role in lateral spikelet development in barley.

Seed storage is important to farmers, breeders and for germplasm preservation. During storage, seeds accumulate damage at the structural and metabolic level, which disrupt their function and reduce resistance to adverse external conditions. In this regard, issues related to seed aging prove to be relevant for maintaining the viability of genetic collections. We analyzed morphological characteristics of grains and their coat color for 44 recombinant inbred lines (RILs) of bread wheat grown in four different seasons, 2003, 2004, 2009 and 2014. Our investigations were performed in 2020. For 19 RILs from the same seasons germination was evaluated. Our results demonstrate that genotype significantly affects the variability of all seed traits, and the year of harvesting affects about 80% of them (including all the traits of shape and size). To identify the trend between changes in grain characteristics and harvesting year, we estimated correlation coefficients between them. No significant trend was detected for the grain shape/size traits, while 90% of the color traits demonstrated such a trend. The most significant negative correlations were found between the harvesting year and the traits of grain redness: the greater the storage time, the more intensive is red color component for the grains. At the same time, it was shown that grains of longer storage time (earlier harvesting year) have lighter coat. Analysis of linear correlations between germination of wheat seeds of different genotypes and harvesting years and their seed traits revealed a negative linear relationship between the red component of coat color and germination: the redder the grains, the lower their germination rate. The results obtained demonstrate manifestations of metabolic changes in the coat of grains associated with storage time and their relationship with a decrease of seed viability.