Chromosome 2HL Research Articles

Genome-wide association mapping for seedling and adult resistance to powdery mildew in barley

Key messageTwo new major QTL were identified for powdery mildew resistance. We confirmed that the QTL on 7HS contributed mainly to the adult-plant resistance, while another one on chromosome arm 1HS made a significant contribution to the seedling resistance.Powdery mildew (PM), caused by Blumeria hordei, can occur at all post emergent stages of barley and constantly threatens crop production. To identify more genes for effective resistance to powdery mildew for use in breeding programs, 696 barley accessions collected from different regions of the world were evaluated for PM resistance at seedling and adult growth stages in three different states of Australia. These barley accessions were genotyped using DArTSeq with over 18,000 markers for a genome-wide association study (GWAS). Using the FarmCPU model, 54 markers showed significant associations with PM resistance scored at the seedling and adult-plant stages in different states of Australia. Another 40 markers showed tentative associations (LOD > 4.0) with resistance. These markers are distributed across all seven barley chromosomes. Most of them were grouped into eleven QTL regions, coinciding with the locations of most of the reported resistance genes. Two major MTAs were identified on chromosome arms 3HS and 5HL, with one on 3HS contributing to adult plant resistance and the one on 5HL to both seedling and adult plant resistance. An MTA on 7HS contributed mainly to the adult-plant resistance, while another one on chromosome arm 1HS made a significant contribution to the seedling resistance.

Telomere and subtelomere high polymorphism might contribute to the specificity of homologous recognition and pairing during meiosis in barley in the context of breeding

Barley (Hordeum vulgare) is one of the most popular cereal crops globally. Although it is a diploid species, (2n = 2x = 14) the study of its genome organization is necessary in the framework of plant breeding since barley is often used in crosses with other cereals like wheat to provide them with advantageous characters. We already have an extensive knowledge on different stages of the meiosis, the cell division to generate the gametes in species with sexual reproduction, such as the formation of the synaptonemal complex, recombination, and chromosome segregation. But meiosis really starts with the identification of homologous chromosomes and pairing initiation, and it is still unclear how chromosomes exactly choose a partner to appropriately pair for additional recombination and segregation. In this work we present an exhaustive molecular analysis of both telomeres and subtelomeres of barley chromosome arms 2H-L, 3H-L and 5H-L. As expected, the analysis of multiple features, including transposable elements, repeats, GC content, predicted CpG islands, recombination hotspots, G4 quadruplexes, genes and targeted sequence motifs for key DNA-binding proteins, revealed a high degree of variability both in telomeres and subtelomeres. The molecular basis for the specificity of homologous recognition and pairing occurring in the early chromosomal interactions at the start of meiosis in barley may be provided by these polymorphisms. A more relevant role of telomeres and most distal part of subtelomeres is suggested.

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.

New introgressive forms of cultivated barley obtained on the basis of interspecific hybrids <i>Hordeum vulgare</i> L. × <i>Hordeum bulbosum</i> L.

Background. The use of alien genetic material of bulbous barley Hordeum bulbosum L. to increase the diversity of cultivated barley Hordeum vulgare L. is an important task, since H. bulbosum is characterized by a number of valuable traits. One of the ways to use the genetic potential of bulbous barley is the interspecific hybridization and obtaining fertile introgressive lines of H. vulgare based on interspecific hybrids. The aim of the study was to obtain new introgressive forms of spring barley using interspecific triploid (HvHbHb) and tetraploid (HbHbHvHv) hybrids of cultivated barley with bulbous barley to expand the collection of introgressive lines of H. vulgare. Materials and methods. To create new introgressive forms, diploid barley H. vulgare (2x) cv. ‘Roland’, was crossed with interspecific hybrids H. vulgare cv. ‘Roland’(2x) × H. bulbosum W851 (4х) (HvHbHb), and H. bulbosum A17 (4x) × H. vulgare ‘Borwina’ (4x) (HbHbHvHv). Cultivated barley forms with introgression of the bulbous barley genetic material were selected from the offspring from these crosses; then the selection was continued in two progenies from self-pollination of selected BC1 plants with three alien introgressions. Identification and localization of introgressions was carried out using the method of fluorescent in situ hybridization (GISH and FISH with chromosome-specific markers). Results. The crossing of cultivated barley with triploid and tetraploid interspecific hybrids H. vulgare × H. bulbosum yielded new forms of cultivated barley with recombinant chromosomes, among which two plants with three terminal introgressions of the genetic material of bulbous barley were identified. The first plant, derived from a triploid hybrid, showed introgressions in chromosomes 5HL, 1HL and 3HS. When two homologues with the 5HL introgression of the initial size are combined in the karyotype, the lethality of seedlings is observed in the offspring plants. In the second plant obtained on the basis of a tetraploid hybrid introgression was revealed in chromosomes 5HL, 2HL, and 7HS. In the offspring from self-pollination of this form, the presence of the 2HL introgression of the original size in both homologues led to plant sterility. Forms with a change in size of the introgression in 5HL and 3HS in the offspring of the first plant and with a change in size of the introgression in 2HL in the offspring of the second plant were detected, which indicated that meiotic recombination had occurred in those chromosome regions in BC1 plants. Conclusions A barley plant with the introgression of bulbous barley chromatin into chromosomes 1HL, 5HL, and 3HS of cultivated barley was identified in the offspring from a cross of cultivated barley with a triploid interspecific hybrid H. vulgare × H. bulbosum. In crosses with a tetraploid interspecific hybrid, a barley plant with the introgression into chromosomes 2HL, 5HL, and 7HS was found. On the basis of ‘Roland’ spring barley cultivar, two series of new introgressive forms of H. vulgare with various combinations of recombinant chromosomes have been created.

Adult resistance genes to barley powdery mildew confer basal penetration resistance associated with broad-spectrum resistance.

Powdery mildew isa major disease of barley (Hordeum vulgare L.) for which breeders have traditionally relied on dominant, pathogen race-specific resistance genes for genetic control. Directional selection pressures in extensive monocultures invariably result in such genes being overcome as the pathogen mutates to evade recognition. This has led to a widespread reliance on fungicides and a single broad-spectrum recessive resistance provided by the mlo gene. The range of resistance genes and alleles found in wild crop relatives and landraces has been reduced in agricultural cultivars through an erosion of genetic diversity during domestication and selective breeding. Three novel major-effect adult plant resistance (APR) genes from landraces, designated Resistance to Blumeria graminis f. sp. hordei (Rbgh1 to Rbgh3), were identified in the terminal regions of barley chromosomes 5HL, 7HS, and 1HS, respectively. The phenotype of the new APR genes showed neither pronounced penetration resistance, nor the spontaneous necrosis and mesophyll cell death typical of mlo resistance, nor a whole epidermal cell hypersensitive response, typical of race-specific resistance. Instead, resistance was localized to the site of attempted penetration in an epidermal cell and was associated with cell wall appositions and cytosolic vesicle-like bodies, and lacked strong induction of reactive oxygen species. The APR genes exhibited differences in vesicle-like body sizes, their distribution, and the extent of localized 3,3-diaminobenzidine staining in individual doubled haploid lines. The results revealed a set of unique basal penetration resistance genes that offer opportunities for combining different resistance mechanisms in breeding programs for robust mildew resistance.

Fine mapping of leaf rust resistance gene Rph13 from wild barley.

Fine mapping of the barley leaf rust resistance locus Rph13 on chromosome 3HL facilitates its use in breeding programs through marker-assisted selection. Barley leaf rust (BLR-caused by Puccinia hordei) is a widespread fungal disease that can be effectively controlled by genetic resistance. There is an ongoing need to both diversify and genetically characterise resistance loci to provide effective and durable control given the ongoing threat of rapidly evolving P. hordei populations. Here, we report on the molecular genetic characterisation of the Rph13 locus, originally derived from wild barley and transferred to barley accession Berac (then referred to as PI 531849). The 2017 reference genome of cv. Morex was used as a road map to rapidly narrow both a genetic and physical intervals around the Rph13 resistance locus. Using recombination-based mapping, we narrowed the physical interval to 116.6kb on chromosome 3H in a segregating population of a cross of the Rph13 carrying resistant line PI 531849 with the leaf rust-susceptible cultivar Gus. We identified two nucleotide-binding leucine-rich repeat genes as likely candidates for the Rph13 resistance. Sequences from the candidate genes enabled the development of a KASP marker that distinguished resistant and susceptible progeny and was found to be predictive and useful for MAS.

Uncovering the evolutionary origin of blue anthocyanins in cereal grains.

Functional divergence after gene duplication plays a central role in plant evolution. Among cereals, only Hordeum vulgare (barley), Triticum aestivum (wheat) and Secale cereale (rye) accumulate delphinidin-derived (blue) anthocyanins in the aleurone layer of grains, whereas Oryza sativa (rice), Zea mays (maize) and Sorghum bicolor (sorghum) do not. The underlying genetic basis for this natural occurrence remains elusive. Here, we mapped the barley Blx1 locus involved in blue aleurone to an approximately 1.13Mb genetic interval on chromosome 4HL, thus identifying a trigenic cluster named MbHF35 (containing HvMYB4H, HvMYC4H and HvF35H). Sequence and expression data supported the role of these genes in conferring blue-coloured (blue aleurone) grains. Synteny analyses across monocot species showed that MbHF35 has only evolved within distinct Triticeae lineages, as a result of dispersed gene duplication. Phylogeny analyses revealed a shared evolution pattern for MbHF35 in Triticeae, suggesting that these genes have co-evolved together. We also identified a Pooideae-specific flavonoid 3',5'-hydroxylase (F3'5'H) lineage, termed here Mo_F35H2, which has a higher amino acid similarity with eudicot F3'5'Hs, demonstrating a scenario of convergent evolution. Indeed, selection tests identified 13 amino acid residues in Mo_F35H2 that underwent positive selection, possibly driven by protein thermostablility selection. Furthermore, through the interrogation of barley germplasm there is evidence that HvMYB4H and HvMYC4H have undergone human selection. Collectively, our study favours blue aleurone as a recently evolved trait resulting from environmental adaptation. Our findings provide an evolutionary explanation for the absence of blue anthocyanins in other cereals and highlight the importance of gene functional divergence for plant diversity and environmental adaptation.

Alternative splicing of a barley gene results in an excess-tillering and semi-dwarf mutant.

An excess-tillering semi-dwarf gene Hvhtd was identified from an EMS-induced mutant in barley and alternative splicing results in excess-tillering semi-dwarf traits. Tillering and plant height are important traits determining plant architecture and grain production in cereal crops. This study identified an excess-tillering semi-dwarf mutant (htd) from an EMS-treated barley population. Genetic analysis of the F1, F2, and F2:3 populations showed that a single recessive gene controlled the excess-tillering semi-dwarf in htd. Using BSR-Seq and gene mapping, the Hvhtd gene was delimited within a 1.8Mb interval on chromosome 2HL. Alignment of the RNA-Seq data with the functional genes in the interval identified a gene HORVU2Hr1G098820 with alternative splicing between exon2 and exon3 in the mutant, due to a G to A single-nucleotide substitution at the exon and intron junction. An independent mutant with a similar phenotype confirmed the result, with alternative splicing between exon3 and exon4. In both cases, the alternative splicing resulted in a non-functional protein. And the gene HORVU2Hr1G098820 encodes a trypsin family protein and may be involved in the IAA signaling pathway and differs from the mechanism of Green Revolution genes in the gibberellic acid metabolic pathway.

Analysis of early-flowering genes at barley chromosome 2H expands the repertoire of mutant alleles at the Mat-c locus

Key messageAnalyses of barley mat-c loss of function mutants reveal deletions, splice-site mutations and nonsynonymous substitutions in a key gene regulating early flowering.Optimal timing of flowering is critical for reproductive success and crop yield improvement. Several major quantitative trait loci for flowering time variation have been identified in barley. In the present study, we analyzed two near-isogenic lines, BW507 and BW508, which were reported to carry two independent early-flowering mutant loci, mat-b.7 and mat-c.19, respectively. Both introgression segments are co-localized in the pericentromeric region of chromosome 2H. We mapped the mutation in BW507 to a 31 Mbp interval on chromosome 2HL and concluded that BW507 has a deletion of Mat-c, which is an ortholog of Antirrhinum majus CENTRORADIALIS (AmCEN) and Arabidopsis thaliana TERMINAL FLOWER1 (AtTFL1). Contrary to previous reports, our data showed that both BW507 and BW508 are Mat-c deficient and none of them are mat-b.7 derived. This work complements previous studies by identifying the uncharacterized mat-c.19 mutant and seven additional mat-c mutants. Moreover, we explored the X-ray structure of AtTFL1 for prediction of the functional effects of nonsynonymous substitutions caused by mutations in Mat-c.

Purple-grained barley (Hordeum vulgare L.): marker-assisted development of NILs for investigating peculiarities of the anthocyanin biosynthesis regulatory network

BackgroundAnthocyanins are plants secondary metabolites important for plant adaptation to severe environments and potentially beneficial to human health. Purple colour of barley grain is caused by the pigments synthesized in pericarp. One or two genes determine the trait. One of them is Ant2 mapped on chromosome 2HL and is known to encode transcription factor (TF) with a bHLH domain. In plants, bHLH regulates anthocyanin biosynthesis together with TF harboring an R2R3-MYB domain. In wheat, the R2R3-MYBs responsible for purple colour of grain pericarp are encoded by the homoallelic series of the Pp-1 genes that were mapped on the short arms of chromosomes 7. In barley, in orthologous positions to wheat’s Pp-1, the Ant1 gene determining red colour of leaf sheath has been mapped. In the current study, we tested whether Ant1 has pleiotropic effect not only on leaf sheath colour but also on pericarp pigmentation.ResultsА set of near isogenic lines (NILs) carrying different combinations of alleles at the Ant1 and Ant2 loci was created using markers-assisted backcrossing approach. The dominant alleles of both the Ant1 and Ant2 genes are required for anthocyanin accumulation in pericarp. A qRT-PCR analysis of the Ant genes in lemma and pericarp of the NILs revealed that some reciprocal interaction occurs between the genes. Expression of each of the two genes was up-regulated in purple-grained line with dominant alleles at the both loci. The lines carrying dominant allele either in the Ant1 or in the Ant2 locus were characterized by the decreased level of expression of the dominant gene and scant activity of the recessive one. The Ant1 and Ant2 expression was barely detected in uncolored line with recessive alleles at both loci. The anthocyanin biosynthesis structural genes were differently regulated: Chs, Chi, F3h, Dfr were transcribed in all lines independently on allelic state of the Ant1 and Ant2 genes, whereas F3’h and Ans were activated in presence on dominant alleles of the both regulatory genes.ConclusionsThe R2R3-MYB-encoding counterpart (Ant1) of the regulatory Ant2 gene was determined for the first time. The dominant alleles of both of them are required for activation of anthocyanin synthesis in barley lemma and pericarp. The R2R3-MYB + bHLH complex activates the synthesis via affecting expression of the F3’h and Ans structural genes. In addition, positive regulatory loop between Ant1 and Ant2 was detected. Earlier the interaction between the anthocyanin biosynthesis regulatory genes has been revealed in dicot plant species only. Our data demonstrated that the regulatory mechanism is considered to be more common for plant kingdom than it has been reported so far.

Duplicated flavonoid 3'-hydroxylase and flavonoid 3', 5'-hydroxylase genes in barley genome.

BackgroundAnthocyanin compounds playing multiple biological functions can be synthesized in different parts of barley (Hordeum vulgare L.) plant. The diversity of anthocyanin molecules is related with branching the pathway to alternative ways in which dihydroflavonols may be modified either with the help of flavonoid 3′-hydroxylase (F3′H) or flavonoid 3′,5′-hydroxylase (F3′5′H)—the cytochrome P450-dependent monooxygenases. The F3′H and F3′5′H gene families are among the least studied anthocyanin biosynthesis structural genes in barley. The aim of this study was to identify and characterise duplicated copies of the F3′H and F3′5′H genes in the barley genome.ResultsFour copies of the F3′5′H gene (on chromosomes 4HL, 6HL, 6HS and 7HS) and two copies of the F3′H gene (on chromosomes 1HL and 6HS) were identified in barley genome. These copies have either one or two introns. Amino acid sequences analysis demonstrated the presence of the flavonoid hydroxylase-featured conserved motifs in all copies of the F3′H and F3′5′H genes with the exception of F3′5′H-3 carrying a loss-of-function mutation in a conservative cytochrome P450 domain. It was shown that the divergence between F3′H and F3′5′H genes occurred 129 million years ago (MYA) before the emergence of monocot and dicot plant species. The F3′H copy approximately occurred 80 MYA; the appearance of F3′5′H copies occurred 8, 36 and 91 MYA. qRT-PCR analysis revealed the tissue-specific activity for some copies of the studied genes. The F3′H-1 gene was transcribed in aleurone layer, lemma and pericarp (with an increased level in the coloured pericarp), whereas the F3′H-2 gene was expressed in stems only. The F3′5′H-1 gene was expressed only in the aleurone layer, and in a coloured aleurone its expression was 30-fold higher. The transcriptional activity of F3′5′H-2 was detected in different tissues with significantly higher level in uncoloured genotype in contrast to coloured ones. The F3′5′H-3 gene expressed neither in stems nor in aleurone layer, lemma and pericarp. The F3′5′H-4 gene copy was weakly expressed in all tissues analysed.ConclusionF3′H and F3′5′H-coding genes involved in anthocyanin synthesis in H. vulgare were identified and characterised, from which the copies designated F3′H-1, F3′H-2, F3′5′H-1 and F3′5′H-2 demonstrated tissue-specific expression patterns. Information on these modulators of the anthocyanin biosynthesis pathway can be used in future for manipulation with synthesis of diverse anthocyanin compounds in different parts of barley plant. Finding both the copies with tissue-specific expression and a copy undergoing pseudogenization demonstrated rapid evolutionary events tightly related with functional specialization of the duplicated members of the cytochrome P450-dependent monooxygenases gene families.

Fast mapping of a chlorophyll b synthesis-deficiency gene in barley (Hordeum vulgare L.) via bulked-segregant analysis with reduced-representation sequencing

Bulked-segregant analysis coupled with next-generation sequencing (BSA-seq) has emerged as an efficient tool for genetic mapping of single genes or major quantitative trait loci controlling (agronomic) traits of interest. However, such a mapping-by-sequencing approach usually relies on deep sequencing and advanced statistical methods. Application of BSA-Seq based on construction of reduced-representation libraries and allele frequency analysis permitted anchoring the barley pale-green (pg) gene on chromosome 3HL. With further marker-assisted validation, pg was mapped to a 3.9 Mb physical-map interval. In the pg mutant a complete deletion of chlorophyllide a oxygenase (HvCAO) gene was identified. Because the product of this gene converts Chl a to Chl b, the pg mutant is deficient in Chl b. An independent Chl b-less mutant line M4437_2 carried a nonsynonymous substitution (F263L) in the C domain of HvCAO. The study demonstrates an optimized pooling strategy for fast mapping of agronomically important genes using a segregating population.

Towards the identification of a gene for prostrate tillers in barley (Hordeum vulgare L.).

Tiller angle, an important agronomic trait, contributes to crop production and plays a vital role in breeding for plant architecture. A barley line V-V-HD, which has prostrate tillers during vegetative growth and erect tillers after booting, is considered the ideal type for repressing weed growth and increasing leaf area during early growth. Genetic analysis identified that the prostrate trait in V-V-HD is controlled by a single gene. A double haploid population with 208 lines from V-V-HD × Buloke was used to map the prostrate growth gene. Ninety-six SNP markers were used for primary mapping, and subsequently, SSR and InDel markers were used for fine mapping. The gene was fine-mapped to a 3.53 Mb region on chromosome 3HL between the markers InDelz3028 and InDelz3032 with 52 candidate genes located in this region. Gene annotation analysis of the 52 genes within the target region indicated that a gene involved in zinc-ion binding (gene ID HORVU3Hr1G090910) is likely to be the candidate gene for prostrate growth in V-V-HD, and is linked to the denso/sdw gene. Association analysis showed that prostrate plants were shorter, flowered later.

Identification and characterization of regulatory network components for anthocyanin synthesis in barley aleurone

BackgroundAmong natural populations, there are different colours of barley (Hordeum vulgare L.). The colour of barley grains is directly related to the accumulation of different pigments in the aleurone layer, pericarp and lemma. Blue grain colour is due to the accumulation of anthocyanins in the aleurone layer, which is dependent on the presence of five Blx genes that are not sequenced yet (Blx1, Blx3 and Blx4 genes clustering on chromosome 4HL and Blx2 and Blx5 on 7HL). Due to the health benefits of anthocyanins, blue-grained barley can be considered as a source of dietary food. The goal of the current study was to identify and characterize components of the anthocyanin synthesis regulatory network for the aleurone layer in barley.ResultsThe candidate genes for components of the regulatory complex MBW (consisting of transcription factors MYB, bHLH/MYC and WD40) for anthocyanin synthesis in barley aleurone were identified. These genes were designated HvMyc2 (4HL), HvMpc2 (4HL), and HvWD40 (6HL). HvMyc2 was expressed in aleurone cells only. A loss-of-function (frame shift) mutation in HvMyc2 of non-coloured compared to blue-grained barley was revealed. Unlike aleurone-specific HvMyc2, the HvMpc2 gene was expressed in different tissues; however, its activity was not detected in non-coloured aleurone in contrast to a coloured aleurone, and allele-specific mutations in its promoter region were found. The single-copy gene HvWD40, which encodes the required component of the regulatory MBW complex, was expressed constantly in coloured and non-coloured tissues and had no allelic differences. HvMyc2 and HvMpc2 were genetically mapped using allele-specific developed CAPS markers developed. HvMyc2 was mapped in position between SSR loci XGBS0875-4H (3.4 cM distal) and XGBM1048-4H (3.4 cM proximal) matching the region chromosome 4HL where the Blx-cluster was found. In this position, one of the anthocyanin biosynthesis structural genes (HvF3’5’H) was also mapped using an allele-specific CAPS-marker developed in the current study.ConclusionsThe genes involved in anthocyanin synthesis in the barley aleurone layer were identified and characterized, including components of the regulatory complex MBW, from which the MYC-encoding gene (HvMyc2) appeared to be the main factor underlying variation of barley by aleurone colour.

Inheritance and characterization of the new and rare gene Rph25 conferring seedling resistance in Hordeum vulgare against Puccinia hordei

AbstractIn this study, we characterized and mapped a new and rare resistance gene (RphFT) in the Chinese barley variety ‘Fong Tien’. RphFT, a dominant gene, was mapped to chromosome 5HL at a genetic position of 142.1 cM using DArT‐seq markers. The gene was also confirmed to be present in Australian cultivar ‘Yagan’ based on allelic tests, and likely ‘Lockyer’ based on multipathotype tests. The genetic studies also confirmed the presence of Rph12 in Australian cultivar ‘Baudin’. Rph12 is also located on chromosome 5HL close to RphFT, and the two loci were confirmed to be independent. Gene RphFT is of limited breeding value because it is effective to only one pathotype of P. hordei, 220P+ +Rph13 in Australia; nevertheless, it may play a role in controlling leaf rust if used in combination with other Rph genes. The locus symbol Rph25 is recommended for RphFT in accordance with the rules and numbering system of barley gene nomenclature.

Interchromosomal Transfer of Immune Regulation During Infection of Barley with the Powdery Mildew Pathogen.

Powdery mildew pathogens colonize over 9500 plant species, causing critical yield loss. The Ascomycete fungus, Blumeria graminis f. sp. hordei (Bgh), causes powdery mildew disease in barley (Hordeum vulgare L.). Successful infection begins with penetration of host epidermal cells, culminating in haustorial feeding structures, facilitating delivery of fungal effectors to the plant and exchange of nutrients from host to pathogen. We used expression Quantitative Trait Locus (eQTL) analysis to dissect the temporal control of immunity-associated gene expression in a doubled haploid barley population challenged with Bgh. Two highly significant regions possessing trans eQTL were identified near the telomeric ends of chromosomes (Chr) 2HL and 1HS. Within these regions reside diverse resistance loci derived from barley landrace H. laevigatum (MlLa) and H. vulgare cv. Algerian (Mla1), which associate with the altered expression of 961 and 3296 genes during fungal penetration of the host and haustorial development, respectively. Regulatory control of transcript levels for 299 of the 961 genes is reprioritized from MlLa on 2HL to Mla1 on 1HS as infection progresses, with 292 of the 299 alternating the allele responsible for higher expression, including Adaptin Protein-2 subunit μ AP2M and Vesicle Associated Membrane Protein VAMP72 subfamily members VAMP721/722. AP2M mediates effector-triggered immunity (ETI) via endocytosis of plasma membrane receptor components. VAMP721/722 and SNAP33 form a Soluble N-ethylmaleimide-sensitive factor Attachment Protein REceptor (SNARE) complex with SYP121 (PEN1), which is engaged in pathogen associated molecular pattern (PAMP)-triggered immunity via exocytosis. We postulate that genes regulated by alternate chromosomal positions are repurposed as part of a conserved immune complex to respond to different pathogen attack scenarios.

Characterization and genetic mapping of the β-diketone deficient eceriferum-b barley mutant.

The barley eceriferum-b.2 (cer-b.2) mutant produces glossy leaf sheaths and is deficient in the cuticular wax component 14,16-hentriacontanedione. The mutated gene maps to a 1.3-cM interval on chromosome 3HL flanked by the genes MLOC_10972 and MLOC_69561. The cuticular wax coating of leaves and stems in many grass species is responsible for the plants' glaucous appearance. A major component of the wax is a group of β-diketone compounds. The barley eceriferum-b.2 (cer-b.2) mutant produces glossy leaf sheaths and is deficient for the compound 14,16-hentriacontanedione. A linkage analysis based on 708 gametes allowed the gene responsible for the mutant phenotype to be mapped to a 1.3-cM interval on chromosome 3HL flanked by the two genes MLOC_10972 and _69561. The product of the wild type allele may represent a step in the β-diketone synthesis pathway.

Genetic analysis of grain and malt quality in an elite barley population

Quantitative trait loci (QTLs) associated with grain weight, grain width, kernel hardness and malting quality were mapped in a doubled haploid population derived from two elite Australian malting barley varieties, Navigator and Admiral. A total of 30 QTLs for grain weight, grain width and kernel hardness were identified in three environments, and 63 QTLs were identified for ten malting quality traits in two environments. Three malting quality traits, namely β-amylase, diastatic power and apparent attenuation limit, were mainly controlled by a QTL linked to the Bmy1 gene at the distal end of chromosome 4H encoding a β-amylase enzyme. Six other malting quality traits, namely α-amylase, soluble protein, Kolbach index, free amino-acid nitrogen, wort β-glucan and viscosity, had coincident QTL clustered on chromosomes 1HS, 4HS, 7HS and 7HL, which demonstrated the interdependence of these traits. There was a strong association between these malt quality QTL clusters on chromosomes 1HS and 7HL and the major QTL for kernel hardness, suggesting that the use of this trait to enable early selection for malting quality in breeding programs would be feasible. In contrast, the majority of QTLs for hot-water extract were not coincident with those identified for other malt quality traits, which suggested differences in the mechanism controlling this trait. Novel QTLs have been identified for kernel hardness on chromosomes 2HL and 7HL, hot-water extract on 7HL and wort β-glucan on 6HL, and the resulting markers may be useful for marker-assisted selection in breeding programs.

QTL mapping reveals genetic architectures of malting quality between Australian and Canadian malting barley (Hordeum vulgare L.)

Australia and Canada are major exporters of malting barley (Hordeum vulgare L.), with Baudin from Australia and AC Metcalfe from Canada being the benchmark varieties for premium malting quality in the past 10 years. We used the barley doubled haploid population derived from a cross of Baudin and AC Metcalfe to map quantitative trait loci (QTLs) for malting quality. The results revealed different genetic architectures controlling malting quality for the two cultivars. Sixteen QTLs were identified and located on chromosomes 1H, 2H, 5H and 7H. The Australian barley Baudin mainly contributed to the malting quality QTL traits of high diastatic power and high β-glucanase on chromosome 1H, while Canadian barley AC Metcalfe mainly contributed to the QTL traits of high hot water extract, high free amino nitrogen, high α-amylase and low malt yield in chromosome 5HL telomere region. This study demonstrated the potential to breed new barley varieties with superior malting quality by integrating genes from Australian and Canadian malting barley varieties. This paper also provides methods to anchor traditional molecular markers without sequence information, such as amplified fragment length polymorphism markers, into the physical map of barley cv. ‘Morex’.

Genomic Regions Influencing Seminal Root Traits in Barley.

Water availability is a major limiting factor for crop production, making drought adaptation and its many component traits a desirable attribute of plant cultivars. Previous studies in cereal crops indicate that root traits expressed at early plant developmental stages, such as seminal root angle and root number, are associated with water extraction at different depths. Here, we conducted the first study to map seminal root traits in barley ( L.). Using a recently developed high-throughput phenotyping method, a panel of 30 barley genotypes and a doubled-haploid (DH) population (ND24260 × 'Flagship') comprising 330 lines genotyped with diversity array technology (DArT) markers were evaluated for seminal root angle (deviation from vertical) and root number under controlled environmental conditions. A high degree of phenotypic variation was observed in the panel of 30 genotypes: 13.5 to 82.2 and 3.6 to 6.9° for root angle and root number, respectively. A similar range was observed in the DH population: 16.4 to 70.5 and 3.6 to 6.5° for root angle and number, respectively. Seven quantitative trait loci (QTL) for seminal root traits (root angle, two QTL; root number, five QTL) were detected in the DH population. A major QTL influencing both root angle and root number (/) was positioned on chromosome 5HL. Across-species analysis identified 10 common genes underlying root trait QTL in barley, wheat ( L.), and sorghum [ (L.) Moench]. Here, we provide insight into seminal root phenotypes and provide a first look at the genetics controlling these traits in barley.