Novel resistance to the Bymovirus BaMMV established by targeted mutagenesis of the PDIL5-1 susceptibility gene in barley.

The Potyviridae are the largest family of plant-pathogenic viruses. Members of this family are the soil-borne bymoviruses barley yellow mosaic virus (BaYMV) and barley mild mosaic virus (BaMMV), which, upon infection of young winter barley seedlings in autumn, can cause yield losses as high as 50%. Resistance breeding plays a major role in coping with these pathogens. However, some viral strains have overcome the most widely used resistance. Thus, there is a need for novel sources of resistance. In ancient landraces and wild relatives of cultivated barley, alleles of the susceptibility factor PROTEIN DISULFIDE ISOMERASE LIKE 5-1 (PDIL5-1) were identified to confer resistance to all known strains of BaYMV and BaMMV. Although the gene is highly conserved throughout all eukaryotes, barley is thus far the only species for which PDIL5-1-based virus resistance has been reported. Whereas introgression by crossing to the European winter barley breeding pool is tedious, time-consuming and additionally associated with unwanted linkage drag, the present study exemplifies an approach to targeted mutagenesis of two barley cultivars employing CRISPR-associated endonuclease technology to induce site-directed mutations similar to those described for PDIL5-1 alleles that render certain landraces resistant. Homozygous primary mutants were produced in winter barley, and transgene-free homozygous M2 mutants were produced in spring barley. A variety of mutants carrying novel PDIL5-1 alleles were mechanically inoculated with BaMMV, by which all frameshift mutations and certain in-frame mutations were demonstrated to confer resistance to this virus. Under greenhouse conditions, virus-resistant mutants showed no adverse effects in terms of growth and yield.

Mining the Australian Grains Gene Bank for Rust Resistance in Barley.

Global barley production is threatened by plant pathogens, especially the rusts. In this study we used a targeted genotype-by-sequencing (GBS) assisted GWAS approach to identify rust resistance alleles in a collection of 287 genetically distinct diverse barley landraces and historical cultivars available in the Australian Grains Genebank (AGG) and originally sourced from Eastern Europe. The accessions were challenged with seven US-derived cereal rust pathogen races including Puccinia hordei (Ph-leaf rust) race 17VA12C, P. coronata var. hordei (Pch-crown rust) race 91NE9305 and five pathogenically diverse races of P. striiformis f. sp. hordei (Psh-stripe rust) (PSH-33, PSH-48, PSH-54, PSH-72 and PSH-100) and phenotyped quantitatively at the seedling stage. Novel resistance factors were identified on chromosomes 1H, 2H, 4H and 5H in response to Pch, whereas a race-specific QTL on 7HS was identified that was effective only to Psh isolates PSH-72 and PSH-100. A major effect QTL on chromosome 5HL conferred resistance to all Psh races including PSH-72, which is virulent on all 12 stripe rust differential tester lines. The same major effect QTL was also identified in response to leaf rust (17VA12C) suggesting this locus contains several pathogen specific rust resistance genes or the same gene is responsible for both leaf rust and stripe rust resistance. Twelve accessions were highly resistant to both leaf and stripe rust diseases and also carried the 5HL QTL. We subsequently surveyed the physical region at the 5HL locus for across the barley pan genome variation in the presence of known resistance gene candidates and identified a rich source of high confidence protein kinase and antifungal genes in the QTL region.

BED domain-containing NLR from wild barley confers resistance to leaf rust.

Leaf rust, caused by Puccinia hordei, is a devastating fungal disease affecting barley (Hordeum vulgare subsp. vulgare) production globally. Despite the effectiveness of genetic resistance, the deployment of single genes often compromises durability due to the emergence of virulent P. hordei races, prompting the search for new sources of resistance. Here we report on the cloning of Rph15, a resistance gene derived from barley's wild progenitor H. vulgare subsp. spontaneum. We demonstrate using introgression mapping, mutation and complementation that the Rph15 gene from the near-isogenic line (NIL) Bowman + Rph15 (referred to as BW719) encodes a coiled-coil nucleotide-binding leucine-rich repeat (NLR) protein with an integrated Zinc finger BED (ZF-BED) domain. A predicted KASP marker was developed and validated across a collection of Australian cultivars and a series of introgression lines in the Bowman background known to carry the Rph15 resistance. Rph16 from HS-680, another wild barley derived leaf rust resistance gene, was previously mapped to the same genomic region on chromosome 2H and was assumed to be allelic with Rph15 based on genetic studies. Both sequence analysis, race specificity and the identification of a knockout mutant in the HS-680 background suggest that Rph15- and Rph16-mediated resistances are in fact the same and not allelic as previously thought. The cloning of Rph15 now permits efficient gene deployment and the production of resistance gene cassettes for sustained leaf rust control.

High-resolution mapping of Rym14Hb, a wild relative resistance gene to barley yellow mosaic disease.

KEY MESSAGE: We mapped the Rym14Hb resistance locus to barley yellow mosaic disease in a 2Mbp interval. The co-segregating markers will be instrumental for marker-assisted selection in barley breeding. Barley yellow mosaic disease is caused by Barley yellow mosaic virus and Barley mild mosaic virus and leads to severe yield losses in barley (Hordeum vulgare) in Central Europe and East-Asia. Several resistance loci are used in barley breeding. However, cases of resistance-breaking viral strains are known, raising concerns about the durability of those genes. Rym14Hb is a dominant major resistance gene on chromosome 6HS, originating from barley's secondary genepool wild relative Hordeum bulbosum. As such, the resistance mechanism may represent a case of non-host resistance, which could enhance its durability. A susceptible barley variety and a resistant H. bulbosum introgression line were crossed to produce a large F2 mapping population (n = 7500), to compensate for a ten-fold reduction in recombination rate compared to intraspecific barley crosses. After high-throughput genotyping, the Rym14Hb locus was assigned to a 2Mbp telomeric interval on chromosome 6HS. The co-segregating markers developed in this study can be used for marker-assisted introgression of this locus into barley elite germplasm with a minimum of linkage drag.

Delineating the elusive BaMMV resistance gene rym15 in barley by medium-resolution mapping.

Barley mild mosaic virus (BaMMV), transmitted by the soil-borne protist Polymyxa graminis, has a serious impact on winter barley production. Previously, the BaMMV resistance gene rym15 was mapped on chromosome 6HS, but the order of flanking markers was non-collinear between different maps. To resolve the position of the flanking markers and to enable map-based cloning of rym15, two medium-resolution mapping populations Igri (susceptible) × Chikurin Ibaraki 1 (resistant) (I × C) and Chikurin Ibaraki 1 × Uschi (susceptible) (C × U), consisting of 342 and 180 F2 plants, respectively, were developed. Efficiency of the mechanical inoculation of susceptible standards varied from 87.5 to 100% and in F2 populations from 90.56 to 93.23%. Phenotyping of F2 plants and corresponding F3 families revealed segregation ratios of 250 s:92r (I × C, χ2 = 0.659) and 140 s:40r (C × U, χ2 = 0.741), suggesting the presence of a single recessive resistance gene. After screening the parents with the 50 K Infinium chip and anchoring corresponding SNPs to the barley reference genome, 8 KASP assays were developed and used to remap the gene. Newly constructed maps revealed a collinear order of markers, thereby allowing the identification of high throughput flanking markers. This study demonstrates how construction of medium-resolution mapping populations in combination with robust phenotyping can efficiently resolve conflicting marker ordering and reduce the size of the target interval. In the reference genome era and genome-wide genotyping era, medium-resolution mapping will help accelerate candidate gene identification for traits where phenotyping is difficult. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-021-01270-9.

Bymovirus-induced yellow mosaic diseases in barley and wheat: viruses, genetic resistances and functional aspects.

Bymovirus-induced yellow mosaic diseases seriously threaten global production of autumn-sown barley and wheat, which are two of the presently most important crops around the world. Under natural field conditions, the diseases are caused by infection of soil-borne plasmodiophorid Polymyxa graminis-transmitted bymoviruses of the genus Bymovirus of the family Potyviridae. Focusing on barley and wheat, this article summarizes the achievements on taxonomy, geography and host specificity of these disease-conferring viruses, as well as the genetics of resistance in barley, wheat and wild relatives. Moreover, based on recent progress of barley and wheat genomics, germplasm resources and large-scale sequencing, the exploration and isolation of corresponding resistant genes from wheat and barley as well as relatives, no matter what a large and complicated genome is present, are becoming feasible and are discussed. Furthermore, the foreseen advances on cloning of the resistance or susceptibility-encoding genes, which will provide the possibility to explore the functional interaction between host plants and soil-borne viral pathogens, are discussed as well as the benefits for marker-assisted resistance breeding in barley and wheat.

Association genetics studies on frost tolerance in wheat (Triticum aestivum L.) reveal new highly conserved amino acid substitutions in CBF-A3, CBF-A15, VRN3 and PPD1 genes.

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.

A saturated SNP linkage map for the orange wheat blossom midge resistance gene Sm1.

KEY MESSAGE: SNP markers were developed for the OWBM resistance gene Sm1 that will be useful for MAS. The wheat Sm1 region is collinear with an inverted syntenic interval in B. distachyon. Orange wheat blossom midge (OWBM, Sitodiplosis mosellana Géhin) is an important insect pest of wheat (Triticum aestivum) in many growing regions. Sm1 is the only described OWBM resistance gene and is the foundation of managing OWBM through host genetics. Sm1 was previously mapped to wheat chromosome arm 2BS relative to simple sequence repeat (SSR) markers and the dominant, sequence characterized amplified region (SCAR) marker WM1. The objectives of this research were to saturate the Sm1 region with markers, develop improved markers for marker-assisted selection (MAS), and examine the synteny between wheat, Brachypodium distachyon, and rice (Oryza sativa) in the Sm1 region. The present study mapped Sm1 in four populations relative to single nucleotide polymorphisms (SNPs), SSRs, Diversity Array Technology (DArT) markers, single strand conformation polymorphisms (SSCPs), and the SCAR WM1. Numerous high quality SNP assays were designed that mapped near Sm1. BLAST delineated the syntenic intervals in B. distachyon and rice using gene-based SNPs as query sequences. The Sm1 region in wheat was inverted relative to B. distachyon and rice, which suggests a chromosomal rearrangement within the Triticeae lineage. Seven SNPs were tested on a collection of wheat lines known to carry Sm1 and not to carry Sm1. Sm1-flanking SNPs were identified that were useful for predicting the presence or absence of Sm1 based upon haplotype. These SNPs will be a major improvement for MAS of Sm1 in wheat breeding programs.

Genetic analyses of BaMMV/BaYMV resistance in barley accession HOR4224 result in the identification of an allele of the translation initiation factor 4e (Hv-eIF4E) exclusively effective against Barley mild mosaic virus (BaMMV).

KEY MESSAGE: Based on a strategy combining extensive segregation analyses and tests for allelism with allele-specific re-sequencing an Hv-eIF4E allele exclusively effective against BaMMV was identified and closely linked markers for BaYMV resistance were developed. Soil-borne barley yellow mosaic disease is one of the most important diseases of winter barley. In extensive screenings for resistance, accession 'HOR4224' being resistant to three strains of Barley mild mosaic virus (BaMMV-ASL1, BaMMV-Sil, and BaMMV-Teik) and two strains of Barley yellow mosaic virus (BaYMV-1 and BaYMV-2) was identified. Analyses using Bmac29, being to some extent diagnostic for the rym4/5 locus, gave hint to the presence of the susceptibility-encoding allele at this locus. Therefore, 107 DH lines derived from the cross 'HOR4224' × 'HOR10714' (susceptible) were screened for resistance. Genetic analyses revealed an independent inheritance of resistance to BaMMV and BaYMV ([Formula: see text] = 5.58) both encoded by a single gene (BaMMV [Formula: see text] = 0.477; BaYMV [Formula: see text] = 0.770). Although Bmac29 indicated the susceptibility-encoding allele, BaMMV resistance of 'HOR4224' co-localized with rym4/rym5. The BaYMV resistance was mapped to chromosome 5H in the region of rym3. Sequencing of full length cDNA of the Hv-eIF4E gene displayed an already sequenced allele described to be efficient against BaMMV and BaYMV. However, the F1 progenies of crosses involving 'HOR4224' and rym4/rym5 donors were all resistant to BaMMV but susceptible to BaYMV. Therefore, this is the first report of an allele at the rym4/rym5 locus exclusively efficient against BaMMV. Changes in the specificity are due to one non-synonymous amino acid substitution (I118K). Results obtained elucidate that combining extensive segregation analyses and tests for allelism involving different strains of BaMMV/BaYMV in combination with allele-specific re-sequencing is an efficient strategy for gene and allele detection in complex pathosystems.

Comparative QTL analysis of root lesion nematode resistance in barley.

KEY MESSAGE: This study demonstrates for the first time that resistance to different root lesion nematodes ( P. neglectus and P. penetrans ) is controlled by a common QTL. A major resistance QTL ( Rlnnp6H ) has been mapped to chromosome 6H using two independent barley populations. Root lesion nematodes (Pratylenchus spp.) are important pests in cereal production worldwide. We selected two doubled haploid populations of barley (Igri × Franka and Uschi × HHOR 3073) and infected them with Pratylenchus penetrans and Pratylenchus neglectus. Nematode multiplication rates were measured 7 or 10 weeks after infection. In both populations, continuous phenotypic variations for nematode multiplication rates were detected indicating a quantitative inheritance of resistance. In the Igri × Franka population, four P. penetrans resistance QTLs were mapped with 857 molecular markers on four linkage groups (2H, 5H, 6H and 7H). In the Uschi × HHOR 3073 population, eleven resistance QTLs (P. penetrans and P. neglectus) were mapped with 646 molecular markers on linkage groups 1H, 3H, 4H, 5H, 6H and 7H. A major resistance QTL named Rlnnp6H (LOD score 6.42-11.19) with a large phenotypic effect (27.5-36.6 %) for both pests was mapped in both populations to chromosome 6H. Another resistance QTL for both pests was mapped on linkage group 5H (Igri × Franka population). These data provide first evidence for common resistance mechanisms against different root lesion nematode species. The molecular markers are a powerful tool for the selection of resistant barley lines among segregating populations because resistance tests are time consuming and laborious.

Major chromosome rearrangements in intergeneric wheat × rye hybrids in compatible and incompatible crosses detected by GBS read coverage analysis.

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.

Genomics-based high-resolution mapping of the BaMMV/BaYMV resistance gene rym11 in barley (Hordeum vulgare L.).

Soil-borne barley yellow mosaic virus disease, caused by different strains of Barley yellow mosaic virus (BaYMV) and Barley mild mosaic virus (BaMMV), is one of the most important diseases of winter barley (Hordeum vulgare L.) in Europe and East Asia. The recessive resistance gene rym11 located in the centromeric region of chromosome 4HL is effective against all so far known strains of BaMMV and BaYMV in Germany. In order to isolate this gene, a high-resolution mapping population (10,204 meiotic events) has been constructed. F2 plants were screened with co-dominant flanking markers and segmental recombinant inbred lines (RILs) were tested for resistance to BaMMV under growth chamber and field conditions. Tightly linked markers were developed by exploiting (1) publicly available barley EST sequences, (2) employing barley synteny to rice, Brachypodium distachyon and sorghum and (3) using next-generation sequencing data of barley. Using this approach, the genetic interval was efficiently narrowed down from the initial 10.72 % recombination to 0.074 % recombination. A marker co-segregating with rym11 was developed providing the basis for gene isolation and efficient marker-assisted selection.

Nutrients and soil structure influence furovirus infection of wheat.

Soil-borne wheat mosaic virus (SBWMV) and Soil-borne cereal mosaic virus (SBCMV), genus Furovirus, family Virgaviridae, cause significant crop losses in cereals. The viruses are transmitted by the soil-borne plasmodiophorid Polymyxa graminis. Inside P. graminis resting spores, the viruses persist in the soil for long time, which makes the disease difficult to combat. To open up novel possibilities for virus control, we explored the influence of physical and chemical soil properties on infection of wheat with SBWMV and SBCMV. Moreover, we investigated, whether infection rates are influenced by the nutritional state of the plants. Infection rates of susceptible wheat lines were correlated to soil structure parameters and nutrient contents in soil and plants. Our results show that SBWMV and SBCMV infection rates decrease the more water-impermeable the soil is and that virus transmission depends on pH. Moreover, we found that contents of several nutrients in the soil (e.g. phosphorous, magnesium, zinc) and in planta (e.g. nitrogen, carbon, boron, sulfur, calcium) affect SBWMV and SBCMV infection rates. The knowledge generated may help paving the way towards development of a microenvironment-adapted agriculture.

SNP Diversity and Genetic Structure of "Rogosija", an Old Western Balkan Durum Wheat Collection.

Durum wheat landraces represent a genetic resource for the identification and isolation of new valuable genes and alleles, useful to increase the crop adaptability to climate change. Several durum wheat landraces, all denominated "Rogosija", were extensively cultivated in the Western Balkan Peninsula until the first half of the 20th century. Within the conservation program of the Montenegro Plant Gene Bank, these landraces were collected, but without being characterized. The main goal of this study was to estimate the genetic diversity of the "Rogosija collection" consisting of 89 durum accessions, using 17 morphological descriptors and the 25K Illumina single nucleotide polymorphism (SNP) array. The genetic structure analysis of the Rogosija collection showed two distinguished clusters localized in two different Montenegro eco-geographic micro-areas, characterized by continental Mediterranean climate and maritime Mediterranean climate. Data suggest that these clusters could be composed of two different Balkan durum landrace collections evolved in two different eco-geographic micro-areas. Moreover, the origin of Balkan durum landraces is discussed.

Fine mapping and comparative genomics integration of two quantitative trait loci controlling resistance to powdery mildew in a Spanish barley landrace.

The intervals containing two major quantitative trait loci (QTL) from a Spanish barley landrace conferring broad spectrum resistance to Blumeria graminis were subjected to marker saturation. First, all the available information on recently developed marker resources for barley was exploited. Then, a comparative genomic analysis of the QTL regions with other sequenced grass model species was performed. As a result of the first step, 32 new markers were added to the previous map and new flanking markers closer to both QTL were identified. Next, syntenic integration revealed that the barley target regions showed homology with regions on chromosome 6 of rice (Oryza sativa), chromosome 10 of Sorghum bicolor and chromosome 1 of Brachypodium distachyon. A nested insertion of ancestral syntenic blocks on Brachypodium chromosome 1 was confirmed. Based on sequence information of the most likely candidate orthologous genes, 23 new barley unigene-derived markers were developed and mapped within the barley target regions. The assessment of colinearity revealed an inversion on chromosome 7HL of barley compared to the other three grass species, and nearly perfect colinearity on chromosome 7HS. This two-step marker enrichment allowed for the refinement of the two QTL into much smaller intervals. Inspection of all predicted proteins for the barley unigenes identified within the QTL intervals did not reveal the presence of resistance gene candidates. This study demonstrates the usefulness of sequenced genomes for fine mapping and paves the way for the use of these two loci in barley breeding programs.

High-Resolution Mapping of Barley mild mosaic virus Resistance Gene rym15.

Barley yellow mosaic virus (BaYMV) and Barley mild mosaic virus (BaMMV), which are transmitted by the soil-borne plasmodiophorid Polymyxa graminis, cause high yield losses in barley. In previous studies, the recessive BaMMV resistance gene rym15, derived from the Japanese landrace Chikurin Ibaraki 1, was mapped on chromosome 6HS of Hordeum vulgare. In this study, 423 F4 segmental recombinant inbred lines (RILs) were developed from crosses of Chikurin Ibaraki 1 with two BaMMV-susceptible cultivars, Igri (139 RILs) and Uschi (284 RILs). A set of 32 competitive allele-specific PCR (KASP) assays, designed using single nucleotide polymorphisms (SNPs) from the barley 50 K Illumina Infinium iSelect SNP chip, genotyping by sequencing (GBS) and whole-genome sequencing (WGS), was used as a backbone for construction of two high-resolution maps. Using this approach, the target locus was narrowed down to 0.161 cM and 0.036 cM in the Igri × Chikurin Ibaraki 1 (I × C) and Chikurin Ibaraki 1 × Uschi (C × U) populations, respectively. Corresponding physical intervals of 11.3 Mbp and 0.281 Mbp were calculated for I × C and C × U, respectively, according to the Morex v3 genome sequence. In the 0.281 Mbp target region, six high confidence (HC) and two low confidence (LC) genes were identified. Genome assemblies of BaMMV-susceptible cultivars Igri and Golden Promise from the barley pan-genome, and a HiFi assembly of Chikurin Ibaraki 1 together with re-sequencing data for the six HC and two LC genes in susceptible parental cultivar Uschi revealed functional SNPs between resistant and susceptible genotypes only in two of the HC genes. These SNPs are the most promising candidates for the development of functional markers and the two genes represent promising candidates for functional analysis.

Dissection of a grain yield QTL from wild emmer wheat reveals sub-intervals associated with culm length and kernel number.

Genetic diversity in wheat has been depleted due to domestication and modern breeding. Wild relatives are a valuable source for improving drought tolerance in domesticated wheat. A QTL region on chromosome 2BS of wild emmer wheat (Triticum turgidum ssp. dicoccoides), conferring high grain yield under well-watered and water-limited conditions, was transferred to the elite durum wheat cultivar Uzan (T. turgidum ssp. durum) by a marker-assisted backcross breeding approach. The 2B introgression line turned out to be higher yielding but also exhibited negative traits that likely result from trans-, cis-, or linkage drag effects from the wild emmer parent. In this study, the respective 2BS QTL was subjected to fine-mapping, and a set of 17 homozygote recombinants were phenotyped at BC4F5 generation under water-limited and well-watered conditions at an experimental farm in Israel and at a high-throughput phenotyping platform (LemnaTec-129) in Germany. In general, both experimental setups allowed the identification of sub-QTL intervals related to culm length, kernel number, thousand kernel weight, and harvest index. Sub-QTLs for kernel number and harvest index were detected specifically under either drought stress or well-watered conditions, while QTLs for culm length and thousand-kernel weight were detected in both conditions. Although no direct QTL for grain yield was identified, plants with the sub-QTL for kernel number showed a higher grain yield than the recurrent durum cultivar Uzan under well-watered and mild drought stress conditions. We, therefore, suggest that this sub-QTL might be of interest for future breeding purposes.

Precision phenotyping across the life cycle to validate and decipher drought-adaptive QTLs of wild emmer wheat (Triticum turgidum ssp. dicoccoides) introduced into elite wheat varieties.

Drought events or the combination of drought and heat conditions are expected to become more frequent due to global warming, and wheat yields may fall below their long-term average. One way to increase climate-resilience of modern high-yielding varieties is by their genetic improvement with beneficial alleles from crop wild relatives. In the present study, the effect of two beneficial QTLs introgressed from wild emmer wheat and incorporated in the three wheat varieties BarNir, Zahir and Uzan was studied under well-watered conditions and under drought stress using non-destructive High-throughput Phenotyping (HTP) throughout the life cycle in a single pot-experiment. Plants were daily imaged with RGB top and side view cameras and watered automatically. Further, at two time points, the quantum yield of photosystem II was measured with a top view FluorCam. The QTL carrying near isogenic lines (NILs) were compared with their corresponding parents by t-test for all non-invasively obtained traits and for the manually determined agronomic and yield parameters. Data quality of phenotypic traits (repeatability) in the controlled HTP experiment was above 85% throughout the life cycle and at maturity. Drought stress had a strong effect on growth in all wheat genotypes causing biomass reduction from 2% up to 70% at early and late points in the drought period, respectively. At maturity, the drought caused 47-55% decreases in yield-related traits grain weight, straw weight and total biomass and reduced TKW by 10%, while water use efficiency (WUE) increased under drought by 29%. The yield-enhancing effect of the introgressed QTLs under drought conditions that were previously demonstrated under field/screenhouse conditions in Israel, could be mostly confirmed in a greenhouse pot experiment using HTP. Daily precision phenotyping enabled to decipher the mode of action of the QTLs in the different genetic backgrounds throughout the entire wheat life cycle. Daily phenotyping allowed a precise determination of the timing and size of the QTLs effect (s) and further yielded information about which image-derived traits are informative at which developmental stage of wheat during the entire life cycle. Maximum height and estimated biovolume were reached about a week after heading, so experiments that only aim at exploring these traits would not need a longer observation period. To obtain information on different onset and progress of senescence, the CVa curves represented best the ongoing senescence of plants. The QTL on 7A in the BarNir background was found to improve yield under drought by increased biomass growth, a higher photosynthetic performance, a higher WUE and a "stay green effect."

The barley leaf rust resistance gene Rph3 encodes a predicted membrane protein and is induced upon infection by avirulent pathotypes of Puccinia hordei.

Leaf rust, caused by Puccinia hordei, is an economically significant disease of barley, but only a few major resistance genes to P. hordei (Rph) have been cloned. In this study, gene Rph3 was isolated by positional cloning and confirmed by mutational analysis and transgenic complementation. The Rph3 gene, which originated from wild barley and was first introgressed into cultivated Egyptian germplasm, encodes a unique predicted transmembrane resistance protein that differs from all known plant disease resistance proteins at the amino acid sequence level. Genetic profiles of diverse accessions indicated limited genetic diversity in Rph3 in domesticated germplasm, and higher diversity in wild barley from the Eastern Mediterranean region. The Rph3 gene was expressed only in interactions with Rph3-avirulent P. hordei isolates, a phenomenon also observed for transcription activator-like effector-dependent genes known as executors conferring resistance to Xanthomonas spp. Like known transmembrane executors such as Bs3 and Xa7, heterologous expression of Rph3 in N. benthamiana induced a cell death response. The isolation of Rph3 highlights convergent evolutionary processes in diverse plant-pathogen interaction systems, where similar defence mechanisms evolved independently in monocots and dicots.

Genome wide association study of frost tolerance in wheat.

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.