Barley Chromosome Research Articles

Characterization of genes required for both Rpg1 and rpg4-mediated wheat stem rust resistance in barley

BackgroundPuccinia graminis f. sp. tritici (Pgt) race TTKSK and its lineage pose a threat to barley production world-wide justifying the extensive efforts to identify, clone, and characterize the rpg4-mediated resistance locus (RMRL), the only effective resistance to virulent Pgt races in the TTKSK lineage. The RMRL contains two nucleotide-binding domain and leucine-rich repeat (NLR) resistance genes, Rpg5 and HvRga1, which are required for resistance. The two NLRs have head-to-head genome architecture with one NLR, Rpg5, containing an integrated C-terminal protein kinase domain, characteristic of an “integrated sensory domain” resistance mechanism. Fast neutron mutagenesis of line Q21861 was utilized in a forward genetics approach to identify genetic components that function in the RMRL or Rpg1 resistance mechanisms, as Q21861 contains both genes. A mutant was identified that compromises both RMRL and Rpg1-mediated resistances and had stunted seedling roots, designated required for P. graminis resistance 9 (rpr9).ResultsThe rpr9 mutant generated in the Q21861 background was crossed with the Swiss landrace Hv584, which carries RMRL but contains polymorphism across the genome compared to Q21861. To map Rpr9, a Hv584 x rpr9 F6:7 recombinant inbred line (RIL) population was developed. The RIL population was phenotyped with Pgt race QCCJB. The Hv584 x rpr9 RIL population was genotyped with the 9 k Illumina Infinium iSelect marker panel, producing 2701 polymorphic markers. A robust genetic map consisting of 563 noncosegregating markers was generated and used to map Rpr9 to an ~ 3.4 cM region on barley chromosome 3H. The NimbleGen barley exome capture array was utilized to capture rpr9 and wild type Q21861 exons, followed by Illumina sequencing. Comparative analysis, resulting in the identification of a 1.05 Mbp deletion at the chromosome 3H rpr9 locus. The identified deletion contains ten high confidence annotated genes with the best rpr9 candidates encoding a SKP1-like 9 protein and a F-box family protein.ConclusionGenetic mapping and exome capture rapidly identified candidate gene/s that function in RMRL and Rpg1 mediated resistance pathway/s. One or more of the identified candidate rpr9 genes are essential in the only two known effective stem rust resistance mechanisms, present in domesticated barley.

Genome-Wide Identification, Characterization and Expression Analysis of Xyloglucan Endotransglucosylase/Hydrolase Genes Family in Barley (Hordeum vulgare).

Xyloglucan endotransglucosylase/hydrolases (XTHs)—a family of xyloglucan modifying enzymes—play an essential role in the construction and restructuring of xyloglucan cross-links. However, no comprehensive study has been performed on this gene family in barley. A total of 24 HvXTH genes (named HvXTH1-24) and an EG16 member were identified using the recently completed genomic database of barley (Hordeum vulgare). Phylogenetic analysis showed that 24 HvXTH genes could be classified into three phylogenetic groups: (I/II, III-A and III-B) and HvXTH15 was in the ancestral group. All HvXTH protein members—except HvXTH15—had a conserved N-glycosylation site. The genomic location of HvXTHs on barley chromosomes showed that the 24 genes are unevenly distributed on the 7 chromosomes, with 10 of them specifically located on chromosome 7H. A structure-based sequence alignment demonstrates that each XTH possesses a highly conserved domain (ExDxE) responsible for catalytic activity. Expression profiles based on the barley genome database showed that HvXTH family members display different expression patterns in different tissues and at different stages. This study is the first systematic genomic analysis of the barley HvXTH gene family. Our results provide valuable information that will help to elucidate the roles of HvXTH genes in the growth and development of barley.

Composition and random elimination of paternal chromosomes in a large population of wheat\u2009\xd7\u2009barley (Triticum aestivum L. \xd7 Hordeum vulgare L.) hybrids

Key messageStatistical analysis of the chromosomal composition in a population of 210 primary plants regenerated from two intergeneric wheat–barley cross combinations revealed the random nature of uniparental elimination for barley chromosomes.Uniparental chromosome elimination is a common process in interspecific and intergeneric cereal hybrids. To characterize the frequency of paternal chromosomes, a population of 218 independent green plants was generated from two wheat (♀) × barley (♂) cross combinations via embryo rescue. The chromosomal composition of 210 primary plants was analyzed with chromosome-specific DNA markers representing all seven barley chromosomes. The analysis revealed an equal proportion of haploid and full hybrids (20.5% and 19.5%, respectively), while the rest of the population contained hypoploids (partial hybrids) with no preference for any possible numbers (one to six) of barley chromosome additions. Contrary to the previous reports, there was no statistical bias or preferential elimination for any individual barley chromosome (1H–7H) in this population. The reasons for the apparent contradiction and the implications of the above findings for cereal breeding are discussed.

Genome-wide meta-analysis on yield and yield-related QTLs in barley (Hordeum vulgare L.)

Meta-QTL (MQTL) analysis improves genetic dissection and detection of consistent chromosomal regions associated with economically important quantitative traits such as yield and yield-related traits through integrating QTL data from independent experiments. In order to identify the most reliable and stable QTLs for grain yield, thousand grain weight, plant height, heading date, harvest index, and number of ears per m2 in barley under normal conditions, we carried out a MQTL analysis on 639 QTLs reported in the last 15 years representing 26 independent mapping populations. Consequently, all the QTLs were condensed to 31 MQTLs distributed on all seven barley chromosomes with a median number of nine initial QTLs and reduced confidence interval mainly to < 2 cM. The QTL and MQTL projection on the barley genome indicated a sub-telomeric enrichment of QTLs likely due to higher gene density and recombination rate at these regions. Finally, the most promising candidate genes (CGs) located in MQTL intervals were presented and discussed based on their revealed function in the orthologous genes. The detected narrowed and stable QTLs and CGs provided a new perspective to improve yield and yield-related traits under normal conditions for breeding programs in barley and its close cereal species.

Fine mapping of Rha2 in barley reveals candidate genes for resistance against cereal cyst nematode

Key messageThe cereal cyst nematode resistance locus Rha2 was mapped to a 978 kbp region on the long arm of barley chromosome 2H. Three candidate genes are discussed.The cereal cyst nematode (CCN) Heterodera avenae is a soil-borne obligate parasite that can cause severe damage to cereals. This research involved fine mapping of Rha2, a CCN resistance locus on chromosome 2H of barley. Rha2 was previously mapped relative to restriction fragment length polymorphisms (RFLPs) in two mapping populations. Anchoring of flanking RFLP clone sequences to the barley genome assembly defined an interval of 5077 kbp. Genotyping-by-sequencing of resistant and susceptible materials led to the discovery of potentially useful single nucleotide polymorphisms (SNPs). Assays were designed for these SNPs and applied to mapping populations. This narrowed the region of interest to 3966 kbp. Further fine mapping was pursued by crossing and backcrossing the resistant cultivar Sloop SA to its susceptible ancestor Sloop. Evaluation of F2 progeny confirmed that the resistance segregates as a single dominant gene. Genotyping of 9003 BC2F2 progeny identified recombinants. Evaluation of recombinant BC2F3 progeny narrowed the region of interest to 978 kbp. Two of the SNPs within this region proved to be diagnostic of CCN resistance across a wide range of barley germplasm. Fluorescence-based and gel-based assays were developed for these SNPs for use in marker-assisted selection. Within the candidate region of the reference genome, there are nine high-confidence predicted genes. Three of these, one that encodes RAR1 (a cysteine- and histidine-rich domain-containing protein), one that is predicted to encode an acetylglutamate kinase and one that is predicted to encode a tonoplast intrinsic protein, are discussed as candidate genes for CCN resistance.

Novel loci and a role for nitric oxide for seed dormancy and preharvest sprouting in barley.

Barley is used for food and feed, and brewing. Nondormant seeds are required for malting, but the lack of dormancy can lead to preharvest sprouting (PHS), which is also undesired. Here, we report several new loci that modulate barley seed dormancy and PHS. Using genome-wide association mapping of 184 spring barley genotypes, we identified four new, highly significant associations on chromosomes 1H, 3H, and 5H previously not associated with barley seed dormancy or PHS. A total of 71 responsible genes were found mostly related to flowering time and hormone signalling. A homolog of the well-known Arabidopsis Delay of Germination 1 (DOG1) gene was annotated on the barley chromosome 3H. Unexpectedly, DOG1 appears to play only a minor role in barley seed dormancy. However, the gibberellin oxidase gene HvGA20ox1 contributed to dormancy alleviation, and another seven important loci changed significantly during after-ripening. Furthermore, nitric oxide release correlated negatively with dormancy and shared 27 associations. Origin and growth environment affected seed dormancy and PHS more than did agronomic traits. Days to anthesis and maturity were shorter when seeds were produced under drier conditions, seeds were less dormant, and PHS increased, with a heritability of 0.57-0.80. The results are expected to be useful for crop improvement.

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

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. 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, β-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. 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.

Pyramiding rpg4- and Rpg1-Mediated Stem Rust Resistance in Barley Requires the Rrr1 Gene for Both to Function.

Stem rust, caused by Puccinia graminis f. sp. tritici (Pgt) is an economically important disease of wheat and barley. Rpg1 is the only resistance gene deployed in Midwestern US barley varieties and provides remarkable resistance to most North American races, except Pgt race QCCJB. Rpg1 is also ineffective against Pgt race TTKSK and its lineage that originated in Africa. The barley rpg4-mediated resistance locus (RMRL) conferring resistance to Pgt races QCCJB and TTKSK was isolated from line Q21861, which is resistant to all known Pgt races due to Rpg1 and RMRL. To develop elite barley varieties RMRL was pyramided into the varieties, Pinnacle and Conlon (both contain Rpg1), producing the near isogenic lines (NILs), Pinnacle RMRL-NIL (PRN) and Conlon RMRL-NIL (CRN). The CRN was resistant to Pgt races QCCJB (RMRL specific) and HKHJC (Rpg1 specific) at the seedling stage and Pgt race TTKSK (RMRL specific) at the adult stage. In contrast, PRN was susceptible to QCCJB and HKHJC at the seedling stage and TTKSK at the adult stage. Interestingly, PRN’s susceptibility to QCCJB and HKHJC showed that RMRL was non-functional in the Pinnacle background but its presence also suppressed Rpg1-mediated resistance. Thus, in the absence of a gene/s found in the Q21861 background, Rpg1 becomes non-functional if RMRL is present, suggesting that another polymorphic gene, that we designated Rrr1 (required for rpg4-mediated resistance 1), is required for RMRL resistance and Rpg1-mediated resistance in the presence of RMRL. Utilizing a PRN/Q21861 derived recombinant inbred line (RIL) population, Rrr1 was delimited to a ∼0.5 MB physical region, slightly proximal (∼1.8 MB) of RMRL on barley chromosome 5H. A second gene, designated required for Rpg1-mediated resistance 2 (Rrr2), with duplicate gene action to Rrr1 in Rpg1-mediated resistance function, was genetically delimited to a physical region of ∼0.7 MB, slightly distal (∼3.1 MB) to Rpg1 on the short arm of barley chromosome 7H. Thus, Rrr1 is required for RMRL resistance and Rrr1 or Rrr2 is required for functional Rpg1-mediated resistance in the presence of the RMRL introgression. Candidate Rrr1 and Rrr2 genes were identified that need to be considered when pyramiding Rpg1 and RMRL in barley.

Development and genetic analysis of wheat double substitution lines carrying Hordeum vulgare 2H and Thinopyrum intermedium 2Ai#2 chromosomes

Thinopyrum intermedium and barley are two close relatives of wheat and carry many genes that are potentially valuable for the improvement of various wheat traits. In this study we created wheat double substitution lines by hybridizing different wheat–Th. intermedium and wheat–barley disomic alien substitution lines, with the aim of using genes in Th. intermedium and barley for wheat breeding and investigating the genetic behavior of alien chromosomes and their wheat homoeologs. As expected, we obtained two types of wheat double substitution lines, 2D2Ai#2(2B)2H(2A) and 2A2Ai#2(2B)2H(2D), in which different group 2 wheat chromosomes were replaced by barley chromosome 2H and Th. intermedium chromosome 2Ai#2. The new materials were characterized using molecular markers, genomic in situ hybridization (GISH), and fluorescent in situ hybridization (FISH). GISH and FISH experiments revealed that the double substitution lines harbor 42 chromosomes including 38 wheat chromosomes, a pair of barley chromosomes, and a pair of Th. intermedium chromosomes. Analysis using specific DNA markers showed that two pairs of wheat homoeologous group 2 chromosomes in the new lines were substituted by a pair of 2H and a pair of 2Ai#2 chromosomes. Chromosome 2H showed a higher transmission rate than 2Ai#2, and both chromosomes were preferentially transmitted between generations via female gametes. Evaluation of botanic and agronomic traits demonstrated that, compared with their parents, the new lines showed similar growth habits and plant type but differences in plant height, flowering date, and self-fertility. Cytological observations using different probes suggested that the double substitution lines showed nearly normal genetic behavior before and during meiosis. The novel substitution lines can potentially be used in wheat meiosis research and breeding programs.

Genetic basis of drought tolerance during seed germination in barley.

Drought is one of the harshest abiotic stresses hindering seed germination, plant growth, and crop productivity. A high rate and uniformity of germination under stressful conditions are vital for crop establishment and growth; thus, for productivity. A better understanding of the genetic architecture of seed germination under drought stress is a prerequisite for further increasing yield potential. Barley is considered one of the most abiotic stresses-tolerant cereals. Elucidating the drought tolerance of barley during seed germination would indeed pave the way towards improving the performance of all cereals. However, we still know relatively little about the genetic control of drought tolerance during the seed germination phase. In our study, 218 worldwide spring barley accessions were subjected to PEG-induced drought during seed germination. Induced drought stress "20% PEG" significantly reduced the seed germination parameters and seedling related traits. A genome-wide association scan (GWAS) was used to identify genomic regions associated with our trait of interest. In total, 338 single nucleotide polymorphisms (SNPs) were found to be associated with several traits distributed across seven barley chromosomes, of which 26 genomic regions were associated with candidate genes. The current study found some of the quantitative trait loci (QTL) that have previously been reported to be linked to seed germination-related traits under drought conditions, as well as some new associations. Noteworthy, the identified QTL colocalized with a number of genes (within interval ±0.5 Mbp) that are exclusively distributed on chromosomes 1H, 2H, and 5H. The annotation of these genes in barley shows their roles in drought tolerance through encoding different transcription factors. The function of the identified genes during seed germination was also confirmed by the annotation of their counterparts in Arabidopsis. The current analyses show the power of the GWAS both for identifying putative candidate genes and for improving plant adaptive traits in barley.

Genome-wide identification, characterization, and evolutionary analysis of NBS-encoding resistance genes in barley.

In this study, a systematic analysis of Nucleotide-Binding Site (NBS) disease resistance (R) gene family in the barley, Hordeum vulgare L. cv. Bowman, genome was performed. Using multiple computational analyses, we could identify 96 regular NBS-encoding genes and characterize them on the bases of structural diversity, conserved protein signatures, genomic distribution, gene duplications, differential expression, selection pressure, codon usage, regulation by microRNAs and phylogenetic relationships. Depending on the presence or absence of CC and LRR domains; the identified NBS genes were assigned to four distinct groups; NBS-LRR (53.1%), CC-NBS-LRR (14.6%), NBS (26%), and CC-NBS (6.3%). NBS-associated domain analysis revealed the presence of signal peptides, zinc fingers, diverse kinases, and other structural features. Eighty-five of the identified NBS-encoding genes were mapped onto the seven barley chromosomes, revealing that 50% of them were located on chromosomes 7H, 2H, and 3H, with a tendency of NBS genes to be clustered in the distal telomeric regions of the barley chromosomes. Nine gene clusters, representing 22.35% of total mapped barley NBS-encoding genes, were found, suggesting that tandem duplication stands for an important mechanism in the expansion of this gene family in barley. Phylogenetic analysis determined 31 HvNBS orthologs from rice and Brachypodium. 87 out of 96 HvNBSs were supported by expression evidence, exhibiting various and quantitatively uneven expression patterns across distinct tissues, organs, and development stages. Fourteen potential miRNA-R gene target pairs were further identified, providing insight into the regulation of NBS genes expression. These findings offer candidate target genes to engineer disease-resistant barley genotypes, and promote our understanding of the evolution of NBS-encoding genes in Poaceae crops.

Fine mapping of a dominant gene conferring resistance to spot blotch caused by a new pathotype of Bipolaris sorokiniana in barley.

We fine-mapped and physically anchored a dominant gene (Rbs7) conferring resistance to spot blotch caused by a new pathotype of Bipolaris sorokiniana in a genomic interval of 304kb on barley chromosome 6H. Spot blotch, caused by Bipolaris sorokiniana, is an economically important disease on barley in the Upper Midwest region of the USA and Prairie Provinces of Canada. A new pathotype (pathotype 7, represented by isolate ND4008) of B. sorokiniana has been identified, which is highly virulent on barley cultivars with resistance to other pathotypes of the fungus. In this study, we fine-mapped a dominant gene conferring resistance to pathotype 7 in the barley line PI 235186. Genetic analysis of the F1 and F2 plants from a cross between PI 356741 (highly susceptible to ND4008) and PI 235186 (highly resistant to ND4008) indicated that a single dominant gene (Rbs7) controls the resistance in PI 235186. This result was confirmed by genetic analysis of the F2:3 families and a recombinant inbred line (RIL) population derived from the same cross. Bulked segregant analysis using simple sequence repeat markers localized Rbs7 on the short arm of chromosome 6H. Additional DNA markers were developed from the 6H pseudomolecule sequence of barley cv. Morex and mapped to the genomic region carrying Rbs7 using the RIL population and F2 recombinants derived from the PI 356741 × PI 235186 cross. Rbs7 was fine-mapped between two markers (M13.06 and M13.37), which spans a physical distance of 304kb on Morex chromosome 6H. These results provide a foundation for future cloning of the resistance gene and development of user-friendly molecular markers that can be used for development of spot-blotch-resistant cultivars in barley breeding programs.

A simple and effective ND-FISH probe design for identifying barley (Hordeum vulgare) chromosomes

Fluorescence in situ hybridization (FISH) has been widely used to analyze the physical mapping of chromosomes and study the chromosome rearrangements during evolution. The conventional FISH technique is time-consuming and labor-intensive as it involves many experimental steps including (1) PCR amplification of the target sequence, (2) recovery of the amplified product, (3) plasmid extraction, (4) fluorescence labeling, as well as (5) denaturing of chromosomes and probes. Here we report an effective method for the design of oligonucleotide probes and the development of a new oligonucleotide probe for non-denaturing FISH in barley. The probe Oligo-442A01 with 6133 copies in barley genome could be mapped to the subtelomeres of barley. However, there were significant differences in signal positions and strengths between Oligo-442A01 and the contrast probe Oligo-HvT01.1, especially on chromosome 3H. These oligonucleotide probes combinations (AGG)5 and Oligo-442A01, (CTT)5 and Oligo-HvT01.1 could effectively differentiate the barley chromosomes.

Genetic mapping of a barley leaf rust resistance gene Rph26 introgressed from Hordeum bulbosum.

The quantitative barley leaf rust resistance gene, Rph26, was fine mapped within a H. bulbosum introgression on barley chromosome 1HL. This provides the tools for pyramiding with other resistance genes. A novel quantitative resistance gene, Rph26, effective against barley leaf rust (Puccinia hordei) was introgressed from Hordeum bulbosum into the barley (Hordeum vulgare) cultivar 'Emir'. The effect of Rph26 was to reduce the observed symptoms of leaf rust infection (uredinium number and infection type). In addition, this resistance also increased the fungal latency period and reduced the fungal biomass within infected leaves. The resulting introgression line 200A12, containing Rph26, was backcrossed to its barley parental cultivar 'Emir' to create an F2 population focused on detecting interspecific recombination within the introgressed segment. A total of 1368 individuals from this F2 population were genotyped with flanking markers at either end of the 1HL introgression, resulting in the identification of 19 genotypes, which had undergone interspecific recombination within the original introgression. F3 seeds that were homozygous for the introgressions of reduced size were selected from each F2 recombinant and were used for subsequent genotyping and phenotyping. Rph26 was genetically mapped to the proximal end of the introgressed segment located at the distal end of chromosome 1HL. Molecular markers closely linked to Rph26 were identified and will enable this disease resistance gene to be combined with other sources of quantitative resistance to maximize the effectiveness and durability of leaf rust resistance in barley breeding. Heterozygous genotypes containing a single copy of Rph26 had an intermediate phenotype when compared with the homozygous resistant and susceptible genotypes, indicating an incompletely dominant inheritance.

Compensation Ability between the Chromosomes of Homoeologous Group 7 of Triticum aestivum L. and Hordeum marinum ssp. gussoneanum Hudson (2n = 28) and Analysis of the Transmission Frequency of Alien 7H1Lmar Chromosome through Gametes in the Progeny of Wheat–Barley Substitution Lines

On the basis of directed chromosome substitution, ditelosomic (DT) wheat–barley substitution lines (2n = 42 = 40 + 2t) with participation of wild tetraploid barley Hordeum marinum ssp. gussoneanum Hudson (2n = 28) were first obtained and analyzed. GISH analysis of the substitution lines showed that they carried a pair of telocentric barley chromosomes. With the help of ditelosomic analysis, the lines were identified as DT7HILmar(7A) and DT7HILmar(7B). On the basis of the compensation test and the study of meiotic stability of substitution lines, it was demonstrated that 7HILmar chromosome of H. marinum was homoeologous to the group 7 chromosomes of common wheat. Analysis of the gamete transfer frequency in the progeny of reciprocal hybrids between F1 7A-7HILmar, 7B-7HILmar, 7D-7HILmar and Saratovskaya 29 cultivar showed that the gamete with alien chromosome n = 20 + t successfully competed with euploid gamete n = 21 in both macrosporogenesis and microsporogenesis.

A locus on barley chromosome 5H affects adult plant resistance to powdery mildew

Adult plant resistance against plant pathogens is of interest as a means to achieve durable resistance. Prior to this research, the barley lines CLE210 (from Uruguay) and Denar (from the Czech Republic) had been reported to exhibit adult-plant resistance against powdery mildew. Here, populations of doubled haploid lines from crosses of these lines with the susceptible cultivar Baudin were evaluated for powdery mildew resistance in field experiments. Using linkage maps constructed from genotyping-by-sequencing (GBS) data, it was determined that differences in resistance were largely attributable to a region on the long arm of chromosome 5H (5HL). Therefore, KASP™ assays were developed based on GBS tag sequences mapped on that chromosome, providing more reliable genetic maps. In each population, a large-effect QTL was mapped on 5HL. As no sequence variation was detected between CLE210 and Denar in this region of 5HL, the two sources of resistance may be identical by descent in the QTL region and carry the same resistance gene. Marker assays from the QTL region were evaluated on a panel of barley lines, providing information that breeders could use to select assays for use in marker-assisted selection.

A Pathogen-Responsive Leucine Rich Receptor Like Kinase Contributes to Fusarium Resistance in Cereals.

Receptor-like kinases form the largest family of receptors in plants and play an important role in recognizing pathogen-associated molecular patterns and modulating the plant immune responses to invasive fungi, including cereal defenses against fungal diseases. But hitherto, none have been shown to modulate the wheat response to the economically important Fusarium head blight (FHB) disease of small-grain cereals. Homologous genes were identified on barley chromosome 6H (HvLRRK-6H) and wheat chromosome 6DL (TaLRRK-6D), which encode the characteristic domains of surface-localized receptor like kinases. Gene expression studies validated that the wheat TaLRRK-6D is highly induced in heads as an early response to both the causal pathogen of FHB disease, Fusarium graminearum, and its’ mycotoxic virulence factor deoxynivalenol. The transcription of other wheat homeologs of this gene, located on chromosomes 6A and 6B, was also up-regulated in response to F. graminearum. Virus-induced gene silencing (VIGS) of the barley HvLRRK-6H compromised leaf defense against F. graminearum. VIGS of TaLRRK-6D in two wheat cultivars, CM82036 (resistant to FHB disease) and cv. Remus (susceptible to FHB), confirmed that TaLRRK-6D contributes to basal resistance to FHB disease in both genotypes. Although the effect of VIGS did not generally reduce grain losses due to FHB, this experiment did reveal that TaLRRK-6D positively contributes to grain development. Further gene expression studies in wheat cv. Remus indicated that VIGS of TaLRRK-6D suppressed the expression of genes involved in salicylic acid signaling, which is a key hormonal pathway involved in defense. Thus, this study provides the first evidence of receptor like kinases as an important component of cereal defense against Fusarium and highlights this gene as a target for enhancing cereal resistance to FHB disease.

Molecular cytogenetic and morphological characterization of two wheat-barley translocation lines.

Barley chromosome 5H, carrying important QTLs for plant adaptation and tolerance to abiotic stresses, is extremely instable in the wheat genetic background and is eliminated in the early generations of wheat-barley crosses. A spontaneous wheat-barley 5HS-7DS.7DL translocation was previously obtained among the progenies of the Mv9kr1 x Igri hybrid. The present work reports on the transfer of the 5HS-7DS.7DL translocation into a modern wheat cultivar, Mv Bodri, in order to use it in the wheat breeding program. The comparison of the hybridization bands of DNA repeats HvT01, pTa71, (GAA)n and the barley centromere-specific (AGGGAG)n in Igri barley and the 5HS-7DS.7DL translocation, together with the visualization of the barley chromatin made it possible to determine the size of the introgressed barley segment, which was approximately 74% of the whole 5HS. Of the 29 newly developed PCR markers, whose source ESTs were selected from the Genome Zipper of barley chromosome 5H, 23 were mapped in the introgressed 1–0.26 FL 5HS bin, three were located in the missing C-0.26 FL region, while three markers were specific for 5HL. The translocation breakpoint was flanked by markers Hv7502 and Hv3949. A comparison of the parental wheat cultivars and the wheat-barley introgression lines indicated that the presence of the translocation improved tillering ability in the Mv9kr1 and Mv Bodri genetic background. The similar or better yield components under high- or low-input cultivation environments, respectively, indicated that the 5HS-7DS.7DL translocation had little or no negative effect on yield components, making it a promising genotype to improve wheat genetic diversity. These results promise to accelerate functional genomic studies on barley chromosome 5H and to support pre-breeding and breeding research on wheat.

Homoeologous Chromosomes From Two Hordeum Species Can Recognize and Associate During Meiosis in Wheat in the Presence of the Ph1 Locus.

Understanding the system of a basic eukaryotic cellular mechanism like meiosis is of fundamental importance in plant biology. Moreover, it is also of great strategic interest in plant breeding since unzipping the mechanism of chromosome specificity/pairing during meiosis will allow its manipulation to introduce genetic variability from related species into a crop. The success of meiosis in a polyploid like wheat strongly depends on regular pairing of homologous (identical) chromosomes and recombination, processes mainly controlled by the Ph1 locus. This means that pairing and recombination of related chromosomes rarely occur in the presence of this locus, making difficult wheat breeding trough the incorporation of genetic variability from related species. In this work, we show that wild and cultivated barley chromosomes associate in the wheat background even in the presence of the Ph1 locus. We have developed double monosomic wheat lines carrying two chromosomes from two barley species for the same and different homoeology chromosome group, respectively. Genetic in situ hybridization revealed that homoeologous Hordeum chromosomes recognize each other and pair during early meiosis in wheat. However, crossing over does not occur at any time and they remained always as univalents during meiosis metaphase I. Our results suggest that the Ph1 locus does not prevent chromosome recognition and pairing but crossing over between homoeologous. The role of subtelomeres in chromosome recognition is also discussed.

Genome-wide association studies of net form of net blotch resistance at seedling and adult plant stages in spring barley collection

Net form of net blotch (NFNB) of barley (Hordeum vulgare L.), caused by Pyrenophora teres f. teres (Ptt) Drechsler (anamorph: Drechslera teres [Sacc.] Shoem.), is considered one of the major constraints of successful barley production in major barley growing regions of the world. Resistance to NFNB was evaluated in a barley collection of 336 genotypes (AM-2014), at seedling stage using isolates LGDPtt.19 and TD10 in the USA, and adult stage in seven hotspot environments in Morocco. The AM-2014 panel was genotyped with 9K SNP markers and genome-wide association studies (GWAS) were carried out using mixed linear model (MLM: Q + K) accounting for population structure (Q) and kinship (K) as covariates. Significant (P < 0.001) marker trait associations were corrected for false discovery rate (FDR) at the q < 0.05. Four genotypes showed an average infection response (IRs ≤ 2) to both isolates, LGDPttt.19 and TD10, at the seedling stage, and 30 genotypes showed resistance in all environments in the field while three genotypes exhibited the highest resistance at both stages. The GWAS of NFNB identified 31 distinct QTLs on all seven barley chromosomes, of which 8 with resistance at seedling stage, 21 were associated with resistance at the adult stage, and two QTLs, QRptt.2H-132.15 and QPtt.6H-54-55, conferred resistance at both stages. Of 31 resistance QTLs reported in this study, 10 QTLs coincided with previously mapped QTL while 21 are novel, thereby validating the GWAS approach used in this study. The resistance sources identified in AM-2014 and QTL mapped in this study are valuable resources for marker-assisted breeding for NFNB resistance in the future.