The barley pan-genome reveals the hidden legacy of mutation breeding.

Genetic diversity is key to crop improvement. Owing to pervasive genomic structural variation, a single reference genome assembly cannot capture the full complement of sequence diversity of a crop species (known as the 'pan-genome'1). Multiple high-quality sequence assemblies are an indispensable component of a pan-genome infrastructure. Barley (Hordeum vulgare L.) is an important cereal crop with a long history of cultivation that is adapted to a wide range of agro-climatic conditions2. Here we report the construction of chromosome-scale sequence assemblies for the genotypes of 20 varieties of barley-comprising landraces, cultivars and a wild barley-that were selected as representatives of global barley diversity. We catalogued genomic presence/absence variants and explored the use of structural variants for quantitative genetic analysis through whole-genome shotgun sequencing of 300 gene bank accessions. We discovered abundant large inversion polymorphisms and analysed in detail two inversions that are frequently found in current elite barley germplasm; one is probably the product of mutation breeding and the other is tightly linked to a locus that is involved in the expansion of geographical range. This first-generation barley pan-genome makes previously hidden genetic variation accessible to genetic studies and breeding.

Root vacuolar Na+ sequestration but not exclusion from uptake correlates with barley salt tolerance.

Soil salinity is a major constraint for the global agricultural production. For many decades, Na+ exclusion from uptake has been the key trait targeted in breeding programs; yet, no major breakthrough in creating salt-tolerant germplasm was achieved. In this work, we have combined the microelectrode ion flux estimation (MIFE) technique for non-invasive ion flux measurements with confocal fluorescence dye imaging technique to screen 45 accessions of barley to reveal the relative contribution of Na+ exclusion from the cytosol to the apoplast and its vacuolar sequestration in the root apex, for the overall salinity stress tolerance. We show that Na+ /H+ antiporter-mediated Na+ extrusion from the root plays a minor role in the overall salt tolerance in barley. At the same time, a strong and positive correlation was found between root vacuolar Na+ sequestration ability and the overall salt tolerance. The inability of salt-sensitive genotypes to sequester Na+ in root vacuoles was in contrast to significantly higher expression levels of both HvNHX1 tonoplast Na+ /H+ antiporters and HvVP1 H+ -pumps compared with tolerant genotypes. These data are interpreted as a failure of sensitive varieties to prevent Na+ back-leak into the cytosol and existence of a futile Na+ cycle at the tonoplast. Taken together, our results demonstrated that root vacuolar Na+ sequestration but not exclusion from uptake played the main role in barley salinity tolerance, and suggested that the focus of the breeding programs should be shifted from targeting genes mediating Na+ exclusion from uptake by roots to more efficient root vacuolar Na+ sequestration.

Multiple horizontal transfers of nuclear ribosomal genes between phylogenetically distinct grass lineages.

The movement of nuclear DNA from one vascular plant species to another in the absence of fertilization is thought to be rare. Here, nonnative rRNA gene [ribosomal DNA (rDNA)] copies were identified in a set of 16 diploid barley (Hordeum) species; their origin was traceable via their internal transcribed spacer (ITS) sequence to five distinct Panicoideae genera, a lineage that split from the Pooideae about 60 Mya. Phylogenetic, cytogenetic, and genomic analyses implied that the nonnative sequences were acquired between 1 and 5 Mya after a series of multiple events, with the result that some current Hordeum sp. individuals harbor up to five different panicoid rDNA units in addition to the native Hordeum rDNA copies. There was no evidence that any of the nonnative rDNA units were transcribed; some showed indications of having been silenced via pseudogenization. A single copy of a Panicum sp. rDNA unit present in H. bogdanii had been interrupted by a native transposable element and was surrounded by about 70 kbp of mostly noncoding sequence of panicoid origin. The data suggest that horizontal gene transfer between vascular plants is not a rare event, that it is not necessarily restricted to one or a few genes only, and that it can be selectively neutral.

Genome-Wide Identification and Characterization of Drought Stress Responsive microRNAs in Tibetan Wild Barley.

Drought stress is a major obstacle to agricultural production. Tibetan wild barley with rich genetic diversity is useful for drought-tolerant improvement of cereals. MicroRNAs (miRNAs) play critical roles in controlling gene expression in response to various environment perturbations in plants. However, the genome-wide expression profiles of miRNAs and their targets in response to drought stress are largely unknown in wild barley. In this study, a polyethylene glycol (PEG) induced drought stress hydroponic experiment was performed, and the expression profiles of miRNAs from the roots of two contrasting Tibetan wild barley genotypes XZ5 (drought-tolerant) and XZ54 (drought-sensitive), and one cultivated barley Tadmor (drought-tolerant) generated by high-throughput sequencing were compared. There were 69 conserved miRNAs and 1574 novel miRNAs in the dataset of three genotypes under control and drought conditions. Among them, seven conserved miRNAs and 36 novel miRNAs showed significantly genotype-specific expression patterns in response to drought stress. And 12 miRNAs were further regarded as drought tolerant associated miRNAs in XZ5, which mostly participate in gene expression, metabolism, signaling and transportation, suggesting that they and their target genes play important roles in plant drought tolerance. This is the first comparation study on the miRNA transcriptome in the roots of two Tibetan wild barley genotypes differing in drought tolerance and one drought tolerant cultivar in response to PEG treatment. Further results revealed the candidate drought tolerant miRNAs and target genes in the miRNA regulation mechanism in wild barley under drought stress. Our findings provide valuable understandings for the functional characterization of miRNAs in drought tolerance.

Convergent Evolution of Effector Protease Recognition by Arabidopsis and Barley.

The Pseudomonas syringae cysteine protease AvrPphB activates the Arabidopsis resistance protein RPS5 by cleaving a second host protein, PBS1. AvrPphB induces defense responses in other plant species, but the genes and mechanisms mediating AvrPphB recognition in those species have not been defined. Here, we show that AvrPphB induces defense responses in diverse barley cultivars. We also show that barley contains two PBS1 orthologs, that their products are cleaved by AvrPphB, and that the barley AvrPphB response maps to a single locus containing a nucleotide-binding leucine-rich repeat (NLR) gene, which we termed AvrPphB Response 1 (Pbr1). Transient coexpression of PBR1 with wild-type AvrPphB but not with a protease inactive mutant triggered defense responses, indicating that PBR1 detects AvrPphB protease activity. Additionally, PBR1 coimmunoprecipitated with barley and Nicotiana benthamiana PBS1 proteins, suggesting mechanistic similarity to detection by RPS5. Lastly, we determined that wheat cultivars also recognize AvrPphB protease activity and contain two putative Pbr1 orthologs. Phylogenetic analyses showed, however, that Pbr1 is not orthologous to RPS5. Our results indicate that the ability to recognize AvrPphB evolved convergently and imply that selection to guard PBS1-like proteins occurs across species. Also, these results suggest that PBS1-based decoys may be used to engineer protease effector recognition-based resistance in barley and wheat.

Genome-wide identification, expression profiles and regulatory network of MAPK cascade gene family in barley.

BACKGROUND: Mitogen-activated protein kinase (MAPK) cascade is a conserved and universal signal transduction module in organisms. Although it has been well characterized in many plants, no systematic analysis has been conducted in barley. RESULTS: Here, we identified 20 MAPKs, 6 MAPKKs and 156 MAPKKKs in barley through a genome-wide search against the updated reference genome. Then, phylogenetic relationship, gene structure and conserved protein motifs organization of them were systematically analyzed and results supported the predictions. Gene duplication analysis revealed that segmental and tandem duplication events contributed to the expansion of barley MAPK cascade genes and the duplicated gene pairs were found to undergone strong purifying selection. Expression profiles of them were further investigated in different organs and under diverse abiotic stresses using the available 173 RNA-seq datasets, and then the tissue-specific and stress-responsive candidates were found. Finally, co-expression regulatory network of MAPK cascade genes was constructed by WGCNA tool, resulting in a complicated network composed of a total of 72 branches containing 46 HvMAPK cascade genes and 46 miRNAs. CONCLUSION: This study provides the targets for further functional study and also contribute to better understand the MAPK cascade regulatory network in barley and beyond.

Managing Fusarium Head Blight in Winter Barley With Cultivar Resistance and Fungicide.

Although there has been research on managing Fusarium head blight (FHB) in spring barley, little has been published on cultivar resistance and optimal fungicide timing for FHB management in winter barley. A 3-year (2015 to 2017) field experiment was conducted to measure FHB resistance of winter barley varieties, gauge the potential benefit from a fungicide, and help determine the optimal timing for fungicide application. The split-plot experiment took place in a misted, inoculated nursery in Raleigh, North Carolina using main plots of four winter barley cultivars (Atlantic, Endeavor, Nomini, and Thoroughbred). Three fungicide treatments were applied to subplots: prothioconazole + tebuconazole at full spike emergence, the same fungicide 6 days later, or no fungicide. The late applications significantly reduced FHB index in each of 3 years and significantly reduced deoxynivalenol (DON) in harvested grain in 2 of the 3 years. Applications at full spike emergence also yielded significant benefit in 1 of the 3 years for each parameter. Neither disease symptoms nor DON gave reason to prefer one of the fungicide timings over the other. Across the 3 years, DON ranked the cultivars Endeavor < Nomini = Thoroughbred < Atlantic. Combining the moderate resistance of Endeavor with a fungicide application and averaging the two timings resulted in a 75% DON reduction compared with unsprayed Atlantic. Taken together, our results indicate that barley growers concerned about minimizing DON should both plant moderately resistant varieties and apply fungicide if there is scab risk. During the same period, 16 commercial winter barley cultivars were tested in from three to seven Virginia and North Carolina environments each, and the DON results were compared after standardization across environments. The winter two-row malting barley cultivars Endeavor and Calypso displayed superior and robust DON resistance across environments.

Association and genome analyses to propose putative candidate genes for malt quality traits.

BACKGROUND: We studied the genetics of nine malt quality traits using association genetics in a panel of North Dakota, ICARDA, and Ethiopian barley lines. Grain samples harvested from Bekoji in 2011 and 2012 were used. RESULTS: The mapping panel revealed strong population structure explained by inflorescence-type, geographic origin, and breeding history. North Dakota germplasm were superior in malt quality traits and they can be donors to improve malt quality properties. We identified 106 marker-trait associations (MTAs) for the nine traits, representing 81 genomic regions across all barley chromosomes. Chromosomes 3H, 5H, and 7H contained most of the MTAs (58.5%). Nearly 18.5% of these genomic regions contained two to three malt quality traits. Within ±250 kb of 81 genomic regions, we recovered 348 barley genes, with some potential impacting malt quality. These include invertase, ß-fructofuranosidase, α-glucosidase, serine carboxypeptidase, and bidirectional sugar transporter SWEET14-like protein. Eighteen of these genes were also previously reported in the Hordeum Toolbox, and 17 of them highly expressed during the germination process. CONCLUSION: The results from this study invite further follow-up functional characterization experiments to relate the genes with individual malt quality traits with higher confidence. It also provides germplasm resources for malt barley improvement. © 2018 Society of Chemical Industry.

A physical, genetic and functional sequence assembly of the barley genome.

Barley (Hordeum vulgare L.) is among the world's earliest domesticated and most important crop plants. It is diploid with a large haploid genome of 5.1 gigabases (Gb). Here we present an integrated and ordered physical, genetic and functional sequence resource that describes the barley gene-space in a structured whole-genome context. We developed a physical map of 4.98 Gb, with more than 3.90 Gb anchored to a high-resolution genetic map. Projecting a deep whole-genome shotgun assembly, complementary DNA and deep RNA sequence data onto this framework supports 79,379 transcript clusters, including 26,159 'high-confidence' genes with homology support from other plant genomes. Abundant alternative splicing, premature termination codons and novel transcriptionally active regions suggest that post-transcriptional processing forms an important regulatory layer. Survey sequences from diverse accessions reveal a landscape of extensive single-nucleotide variation. Our data provide a platform for both genome-assisted research and enabling contemporary crop improvement.

Free and Bound Phenolic Compound Content and Antioxidant Activity of Different Cultivated Blue Highland Barley Varieties from the Qinghai-Tibet Plateau.

In this study, the polyphenols composition and antioxidant properties of 12 blue highland barley varieties planted on the Qinghai-Tibet Plateau area were measured. The contents of the free, bound and total phenolic acids varied between 166.20-237.60, 170.10-240.75 and 336.29-453.94 mg of gallic acid equivalents per 100 g of dry weight (DW) blue highland barley grains, while the free and bound phenolic acids accounted for 50.09% and 49.91% of the total phenolic acids, respectively. The contents of the free, bound and total flavones varied among 20.61-25.59, 14.91-22.38 and 37.91-47.98 mg of catechin equivalents per 100 g of dry weight (DW) of blue highland barley grains, while the free and bound flavones accounted for 55.90% and 44.10% of the total flavones, respectively. The prominent phenolic compounds in the blue hulless barley grains were gallic acid, benzoic acid, syringic acid, 4-coumaric acid, naringenin, hesperidin, rutin, (+)-catechin and quercetin. Among these, protocatechuic acid, chlorogenic acid and (+)-catechin were the major phenolic compounds in the free phenolics extract. The most abundant bound phenolics were gallic acid, benzoic acid, syringic acid, 4-coumaric acid, benzoic acid, dimethoxybenzoic acid, naringenin, hesperidin, quercetin and rutin. The average contribution of the bound phenolic extract to the DPPH• free radical scavenging capacity was higher than 86%, that of free phenolic extract to the ABTS•+ free radical scavenging capacity was higher than 79%, and that of free phenolic (53%) to the FRAP antioxidant activity was equivalent to that of the bound phenol extract (47%). In addition, the planting environment exerts a very important influence on the polyphenol composition, content and antioxidant activity of blue highland barley. The correlation analysis showed that 2,4-hydroxybenzoic acid and protocatechuic acid were the main contributors to the DPPH• and ABTS•+ free radical scavenging capacity in the free phenolic extract, while chlorogenic acid, vanillic acid, ferulic acid and quercetin were the main contributors to the free radical scavenging capacity in the bound phenol extract. The study results show that the blue highland barley grains have rich phenolic compounds and high antioxidant activity, as well as significant varietal differences. The free and bound phenolic extracts in the blue hulless barley grains have an equivalent proportion in the total phenol, and co-exist in two forms. They can be used as a potential valuable source of natural antioxidants, and can aid in enhancing the development and daily consumption of foods relating to blue highland barley.

Variable Levels of Tolerance to Water Stress (Drought) and Associated Biochemical Markers in Tunisian Barley Landraces.

Due to its high tolerance to abiotic stress, barley (Hordeum vulgare) is cultivated in many arid areas of the world. In the present study, we evaluate the tolerance to water stress (drought) in nine accessions of "Ardhaoui" barley landraces from different regions of Tunisia. The genetic diversity of the accessions is evaluated with six SSR markers. Seedlings from the nine accessions are subjected to water stress by completely stopping irrigation for three weeks. A high genetic diversity is detected among the nine accessions, with no relationships between genetic distance and geographical or ecogeographical zone. The analysis of growth parameters and biochemical markers in the water stress-treated plants in comparison to their respective controls indicated great variability among the studied accessions. Accession 2, from El May Island, displayed high tolerance to drought. Increased amounts of proline in water-stressed plants could not be correlated with a better response to drought, as the most tolerant accessions contained lower levels of this osmolyte. A good correlation was established between the reduction of growth and degradation of chlorophylls and increased levels of malondialdehyde and total phenolics. These biochemical markers may be useful for identifying drought tolerant materials in barley.

A microsatellite diversity analysis and the development of core-set germplasm in a large hulless barley (Hordeum vulgare L.) collection.

BACKGROUND: Clarifying genetic diversity in a large germplasm resource plays important roles in experimental designs that provides flexible utility in fundamental research and breeding in crops. However, the work is limited due to small collections of barley that are insufficient representatives. RESULTS: In the present study, we collected 562 hulless barley (Hordeum vulgare L.) accessions with worldwide geographic origins and evaluated their genetic variability and relatedness based on 93 simple sequence repeat (SSR) markers. In an integrated analysis of the population structure, analysis of molecular variance (AMOVA) and pairwise F ST, the 562 barley accessions exhibited a strong stratification that allowed for them to be divided into two major subpopulations (p1 and p2) and an admixture subpopulation, with 93, 408 and 61 accessions, respectively. In a neutral test, considerable proportions of SSR alleles expressed the strong non-neutrality in specific subpopulations (44 and 37), which are probably responsible for population differentiation. To reduce the diversity redundancy in large barley collections, we delicately selected a core set of 200 barley accessions as a tradeoff between diversity and representativeness in an easily handled population. In comparing the 562 barley accessions, the core barley set accounted for 96.2% of allelic diversity and 93% to 95% of phenotypic variability, whereas it exhibited a significant enhancement in minor allelic frequencies, which probably benefit association mapping in the barley core set. CONCLUSIONS: The results provided additional insight into the genetic structure in a large barley germplasm resource, from which an easily manageable barley core set was identified, demonstrating the great potential for discovering key QTLs and ultimately facilitating barley breeding progress.

On the allopolyploid origin and genome structure of the closely related species Hordeum secalinum and Hordeum capense inferred by molecular karyotyping.

Background and Aims: To provide additional information to the many phylogenetic analyses conducted within Hordeum , here the origin and interspecific affinities of the allotetraploids Hordeum secalinum and Hordeum capense were analysed by molecular karyotyping. Methods: Karyotypes were determined using genomic in situ hybridization (GISH) to distinguish the sub-genomes and , plus fluorescence in situ hybridization (FISH)/non-denaturing (ND)-FISH to determine the distribution of ten tandem repetitive DNA sequences and thus provide chromosome markers. Key Results: Each chromosome pair in the six accessions analysed was identified, allowing the establishment of homologous and putative homeologous relationships. The low-level polymorphism observed among the H. secalinum accessions contrasted with the divergence recorded for the sub-genome of the H. capense accessions. Although accession H335 carries an intergenomic translocation, its chromosome structure was indistinguishable from that of H. secalinum . Conclusion: Hordeum secalinum and H. capense accession H335 share a hybrid origin involving Hordeum marinum subsp. gussoneanum as the genome donor and an unidentified genome progenitor. Hordeum capense accession BCC2062 either diverged, with remodelling of the sub-genome, or its genome was donated by a now extinct ancestor. A scheme of probable evolution shows the intricate pattern of relationships among the Hordeum species carrying the genome (including all H. marinum taxa and the hexaploid Hordeum brachyantherum ).

Rapid cultivar identification of barley seeds through disjoint principal component modeling.

Classification of barley varieties is a crucial part of the control and assessment of barley seeds especially for the malting and brewing industry. The correct classification of barley is essential in that a majority of decisions made regarding process specifications, economic considerations, and the type of product produced with the cereal are made based on the barley variety itself. This fact combined with the need to promptly assess the cereal as it is delivered to a malt house or production facility creates the need for a technique to quickly identify a barley variety based on a sample. This work explores the feasibility of differentiating between barley varieties based on the protein spectrum of barley seeds. In order to produce a rapid analysis of the protein composition of the barley seeds, lab-on-a-chip micro fluid technology is used to analyze the protein composition. Classification of the barley variety is then made using disjoint principle component models. This work included 19 different barley varieties. The varieties consisted of both winter and summer barley types. In this work, it is demonstrated that this system can identify the most likely barley variety with an accuracy of 95.9% based on cross validation and can screen summer barley with an accuracy of 95.2% and a false positive rate of 0.0% based on cross validation. This demonstrates the feasibility of the method to provide a rapid and relatively inexpensive method to verify the heritage of barley seeds.

The Effect of Homoeologous Meiotic Pairing in Tetraploid Hordeum bulbosum L. × H. vulgare L. Hybrids on Alien Introgressions in Offspring.

The pairing behaviour of the individual chromosome arms of Hordeum vulgare (Hv) with their homoeologous arms of H. bulbosum (Hb) at metaphase I of meiosis in tetraploid Hb × Hv hybrids and the frequencies of recombined Hv chromosome arms in selfed offspring were studied on differentially visualized chromosomes after fluorescent in situ hybridisation. The frequencies of paired Hv-Hb arms in the F2 and F3 hybrids were correlated with the frequencies of recombined Hv chromosomes in progenies. Self-generation of hybrids, the number of Hv and Hb chromosomes, and the number of recombined Hv chromosomes of the hybrids strongly influenced the Hv-Hb pairing frequency in meiosis. Within the offspring of F2 and F3 hybrids both Hv plants and hybrids were detected. In contrast, all progenies of the F4 hybrid were hybrids which exhibited centromere misdivisions. The highest frequencies of homoeologous pairing in hybrids and most recombinants were obtained for the barley chromosome 1HL. Recombinants for 4HL, 5HS, 6HS, and 7HS were rarely found. Meiotic pairing and recombinants involving chromosome 1HS were never observed. The results of this study demonstrate that fertile tetraploid interspecific hybrids with a high intergenomic pairing at meiosis are valuable basic material for introgression breeding in barley.

Allopolyploidy and the complex phylogenetic relationships within the Hordeum brachyantherum taxon.

Hordeum brachyantherum Nevski includes two subspecies: the diploid (2×) subsp. californicum, and subsp. brachyantherum, which itself includes a tetraploid (4×) and a hexaploid (6×) cytotype. The phylogenetic relationships between these taxa and the origin of the polyploids remain controversial. To provide additional information to the many molecular phylogenetic analyses conducted within Hordeum, FISH-based karyotypes were produced for all subspecies/cytotypes within H. brachyantherum. Chromosomes of H. roshevitzii and H. marinum subsp. gussoneanum were also analysed since these species are potentially involved in the origin of the polyploids. For karyotyping, ten repetitive DNA sequences were screened to indentify repeats showing sufficient diversity in terms of copy number and localisation that they might serve as physical markers for distinguishing between each mitotic chromosome pair in all accessions. Genomic in situ hybridisation (GISH) was used to distinguish between subgenomes in polyploids. The karyotype maps allowed the assessment of the chromosomal diversity within species/cytotypes and the identification of possibly homoeologous chromosomes. The results show a wide divergence between the chromosomes of subsp. californicum and H. roshevitzii, and with their supposed derivatives in subsp. brachyantherum 4×. One of the three subgenomes of subsp. brachyantherum 6× is derived from subsp. gussoneanum with no genomic reorganisation (i.e., neither amplification nor loss of the repetitive DNA sequences analysed). It is generally accepted that subsp. brachyantherum 4× is the other progenitor of subsp. brachyantherum 6×, but the present results suggest this to be unlikely. The present findings thus show the cytogenetic diversity and genomic structure of H. brachyantherum, and reveal its complex evolutionary history, in which chromosomal diversification and allopolyploidy have played important roles.

Structure and expression of phosphoglucan phosphatase genes of Like Sex Four1 and Like Sex Four2 in barley.

Phosphoglucan phosphatases (Like-SEX4 1 and 2; LSF1 and LSF2) were reported to play roles in starch metabolism in leaves of Arabidopsis. In this study, we identified and mapped the LSF1 and LSF2 genes in barley (HvLSF1 and HvLSF2), characterized their gene and protein structures, predicted the cis-elements of their promoters, and analysed their expression patterns. HvLSF1 and HvLSF2 were mapped on the long arm of chromosome 1H (1HL) and 5H (5HL), respectively. Our results revealed varied exon-intron structures and conserved exon-intron junctions in both LSF1 and LSF2 from a range of analysed species. Alignment of protein sequences indicated that cTP and CT domains are much less varied than the functional domains (PDZ, DPS and CBM48). LSF2 was mainly expressed in anthers of barley and rice, and in leaf of Arabidopsis. LSF1 was mainly expressed in endosperm of barley and leaf of Arabidopsis and rice. The expression of LSF1 exhibited a diurnal pattern in rice only and that of LSF2 in both rice and Arabidopsis. Of the investigated stresses, only cold stress significantly reduced expression level of LSF1 and LSF2 in barley and LSF2 in Arabidopsis at late stages of the treatments. While heat treatment significantly decreased expression levels of LSF1 at middle stage (4 h) of a treatment in Arabidopsis only. The strong relationships detected between LSF2 and starch excess4 (SEX4), glucan, water dikinases or phosphoglucan, water dikinases were identified and discussed. Taken together, these results provide information of genetic manipulation of LSF1 and LSF2, especially in monocotyledon and further elucidate their regulatory mechanism in plant development.

Species-Level Phylogeny and Polyploid Relationships in Hordeum (Poaceae) Inferred by Next-Generation Sequencing and In Silico Cloning of Multiple Nuclear Loci.

Polyploidization is an important speciation mechanism in the barley genus Hordeum. To analyze evolutionary changes after allopolyploidization, knowledge of parental relationships is essential. One chloroplast and 12 nuclear single-copy loci were amplified by polymerase chain reaction (PCR) in all Hordeum plus six out-group species. Amplicons from each of 96 individuals were pooled, sheared, labeled with individual-specific barcodes and sequenced in a single run on a 454 platform. Reference sequences were obtained by cloning and Sanger sequencing of all loci for nine supplementary individuals. The 454 reads were assembled into contigs representing the 13 loci and, for polyploids, also homoeologues. Phylogenetic analyses were conducted for all loci separately and for a concatenated data matrix of all loci. For diploid taxa, a Bayesian concordance analysis and a coalescent-based dated species tree was inferred from all gene trees. Chloroplast matK was used to determine the maternal parent in allopolyploid taxa. The relative performance of different multilocus analyses in the presence of incomplete lineage sorting and hybridization was also assessed. The resulting multilocus phylogeny reveals for the first time species phylogeny and progenitor-derivative relationships of all di- and polyploid Hordeum taxa within a single analysis. Our study proves that it is possible to obtain a multilocus species-level phylogeny for di- and polyploid taxa by combining PCR with next-generation sequencing, without cloning and without creating a heavy load of sequence data.

Effects of variety and steeping conditions on some barley components associated with colonic health.

BACKGROUND: Butyric acid is produced by degradation of dietary fibre by microbiota and is crucial for maintaining a healthy colon. The physicochemical properties are important for butyric acid formation, and this study aimed to evaluate the use of malting to tailor the functional characteristics of barley dietary fibre. The effect of different steeping conditions was evaluated in laboratory-scale malting experiments with three different barley varieties. RESULTS: Steeping at 35°C and with 0.4 % (v/v) lactic acid resulted in a higher content of ß-glucan and soluble fibre in malts than in those steeped at lower temperature and lower lactic acid concentration. Resistant starch increased, whereas the content of soluble arabinoxylan was lower. Dietary fibre components in Tipple were more affected by steeping conditions than the other varieties. The total contents of iron, phytate and amylose were little influenced by steeping conditions. CONCLUSION: The selection of steeping conditions during malting influences composition and the characteristics of dietary fibre in barley. However, the choice of barley variety is also important for tailoring of functional ingredients beneficial for colonic health. © 2016 The Authors. Journal of the Science of Food and Agriculture published by JohnWiley & Sons Ltd on behalf of Society of Chemical Industry.

Sr33 & Sr35 gene homolog indentification in genomes of cereals related with Aegilops tauschii & Triticum monococcum.

Using bioinformatics analysis, the homologues of the genes Sr33 and Sr35 were identifed in the genomes of Triticum aestivum, Hordeum vulgare and Triticum urartu. It is known that these genes provide resistance to hightly virulent wheat stem rust races (Ug99). To identify important for resistance amino acid sites, the comparison of the founded homologues with the Sr33 and Sr35 protein sequences was performed. It was found that the sequences S5DMA6 and E9P785 are the closest homologues of RGA1e protein ­ a product of the Sr33 gene, and the sequences M7YFA9 (CNL-C) and F2E9R2 are the homologues of CNL9 ­ a product of the gene Sr35. It is assumed that the homologues of the genes Sr33 and Sr35, which derived from the wild relatives of wheat and barley, can provide resistance to various forms of a stem rust and can be used in the future breeding programs for wheat improvement.