An affordable and convenient diagnostic marker to identify male and female hop plants.

Hop production utilizes exclusively female plants, whereas male plants only serve to generate novel variation within breeding programs through crossing. Currently, hop lacks a rapid and accurate diagnostic marker to determine whether plants are male or female. Without a diagnostic marker, breeding programs may take 1-2 years to determine the sex of new seedlings. Previous research on sex-linked markers was restricted to specific populations or breeding programs and therefore had limited transferability or suffered from low scalability. A large collection of 765 hop genotypes with known sex phenotypes, genotyping-by-sequencing, and genome-wide association mapping revealed a highly significant marker on the sex chromosome (LOD score = 208.7) that predicted sex within our population with 96.2% accuracy. In this study, we developed a PCR allele competitive extension (PACE) assay for the diagnostic SNP and tested three quick DNA extraction methodologies for rapid, high-throughput genotyping. Additionally, the marker was validated in a separate population of 94 individuals from 15 families from the USDA-ARS hop breeding program in Prosser, WA with 96% accuracy. This diagnostic marker is located in a gene predicted to encode the basic helix-loop-helix transcription factor protein, a family of proteins that have been previously implicated in male sterility in a variety of plant species, which may indicate a role in determining hop sex. The marker is diagnostic, accurate, affordable, and highly scalable and has the potential to improve efficiency in hop breeding.

Wild barley (Hordeum spontaneum) and landraces (Hordeum vulgare) from Turkey contain an abundance of novel Rhynchosporium commune resistance loci.

KEY MESSAGE: Rhynchosporium commune is a globally devastating pathogen of barley. Wild and landrace barley are underutilized, however, contain an abundance of loci that can be used as potential sources of resistance. Rhynchosporium commune, the causal agent of the disease scald or leaf blotch of barley, is a hemibiotrophic fungal pathogen of global importance, responsible for yield losses ranging from 30 to 40% on susceptible varieties. To date, over 150 resistance loci have been characterized in barley. However, due to the suspected location of the R. commune host jump in Europe, European germplasm has been the primary source used to screen for R. commune resistance leaving wild (Hordeum spontaneum) and landrace (H. vulgare) barley populations from the center of origin largely underutilized. A diverse population consisting of 94 wild and 188 barley landraces from Turkey were genotyped using PCR-GBS amplicon sequencing and screened with six Turkish R. commune isolates. The isolates were collected from distinct geographic regions of Turkey with two from the Aegean region, two from central Turkey and two from the Fertile Crescent region. The data set was utilized for association mapping analysis with a total of 21 loci identified, of which 12 were novel, indicating that these diverse primary barley gene pools contain an abundance of novel R. commune resistances that could be utilized for resistance breeding.

Association mapping reveals a reciprocal virulence/avirulence locus within diverse US Pyrenophora teres f. maculata isolates.

BACKGROUND: Spot form net blotch (SFNB) caused by the necrotrophic fungal pathogen Pyrenophora teres f. maculata (Ptm) is an economically important disease of barley that also infects wheat. Using genetic analysis to characterize loci in Ptm genomes associated with virulence or avirulence is an important step to identify pathogen effectors that determine compatible (virulent) or incompatible (avirulent) interactions with cereal hosts. Association mapping (AM) is a powerful tool for detecting virulence loci utilizing phenotyping and genotyping data generated for natural populations of plant pathogenic fungi. RESULTS: Restriction-site associated DNA genotyping-by-sequencing (RAD-GBS) was used to generate 4,836 single nucleotide polymorphism (SNP) markers for a natural population of 103 Ptm isolates collected from Idaho, Montana and North Dakota. Association mapping analyses were performed utilizing the genotyping and infection type data generated for each isolate when challenged on barley seedlings of thirty SFNB differential barley lines. A total of 39 marker trait associations (MTAs) were detected across the 20 barley lines corresponding to 30 quantitative trait loci (QTL); 26 novel QTL and four that were previously mapped in Ptm biparental populations. These results using diverse US isolates and barley lines showed numerous barley-Ptm genetic interactions with seven of the 30 Ptm virulence/avirulence loci falling on chromosome 3, suggesting that it is a reservoir of diverse virulence effectors. One of the loci exhibited reciprocal virulence/avirulence with one haplotype predominantly present in isolates collected from Idaho increasing virulence on barley line MXB468 and the alternative haplotype predominantly present in isolates collected from North Dakota and Montana increasing virulence on barley line CI9819. CONCLUSIONS: Association mapping provided novel insight into the host pathogen genetic interactions occurring in the barley-Ptm pathosystem. The analysis suggests that chromosome 3 of Ptm serves as an effector reservoir in concordance with previous reports for Pyrenophora teres f. teres, the causal agent of the closely related disease net form net blotch. Additionally, these analyses identified the first reported case of a reciprocal pathogen virulence locus. However, further investigation of the pathosystem is required to determine if multiple genes or alleles of the same gene are responsible for this genetic phenomenon.

Genome-wide association mapping of Pyrenophora teres f. maculata and Pyrenophora teres f. teres resistance loci utilizing natural Turkish wild and landrace barley populations.

Unimproved landraces and wild relatives of crops are sources of genetic diversity that were lost post domestication in modern breeding programs. To tap into this rich resource, genome-wide association studies in large plant genomes have enabled the rapid genetic characterization of desired traits from natural landrace and wild populations. Wild barley (Hordeum spontaneum), the progenitor of domesticated barley (Hordeum vulgare), is dispersed across Asia and North Africa, and has co-evolved with the ascomycetous fungal pathogens Pyrenophora teres f. teres and P. teres f. maculata, the causal agents of the diseases net form of net blotch and spot form of net blotch, respectively. Thus, these wild and local adapted barley landraces from the region of origin of both the host and pathogen represent a diverse gene pool to identify new sources of resistance, due to millions of years of co-evolution. The barley-P. teres pathosystem is governed by complex genetic interactions with dominant, recessive, and incomplete resistances and susceptibilities, with many isolate-specific interactions. Here, we provide the first genome-wide association study of wild and landrace barley from the Fertile Crescent for resistance to both forms of P. teres. A total of 14 loci, four against P. teres f. maculata and 10 against P. teres f. teres, were identified in both wild and landrace populations, showing that both are genetic reservoirs for novel sources of resistance. We also highlight the importance of using multiple algorithms to both identify and validate additional loci.

A guide to plant TPX2-like and WAVE-DAMPENED2-like proteins.

TPX2 proteins were first identified in vertebrates as a key mitotic spindle assembly factor. Subsequent studies demonstrated that TPX2 is an intricate protein, with functionally and structurally distinct domains and motifs including Aurora kinase-binding, importin-binding, central microtubule-binding, and C-terminal TPX2 conserved domain, among others. The first plant TPX2-like protein, WAVE-DAMPENED2, was identified in Arabidopsis as a dominant mutation responsible for reducing the waviness of roots grown on slanted agar plates. Each plant genome encodes at least one 'canonical' protein with all TPX2 domains and a family of proteins (20 in Arabidopsis) that diversified to contain only some of the domains. Although all plant TPX2-family proteins to date bind microtubules, they function in distinct processes such as cell division, regulation of hypocotyl cell elongation by hormones and light signals, vascular development, or abiotic stress tolerance. Consequently, their expression patterns, regulation, and functions have diverged considerably. Here we summarize the current body of knowledge surrounding plant TPX2-family proteins.

The Barley HvWRKY6 Transcription Factor Is Required for Resistance Against Pyrenophora teres f. teres.

Barley is an important cereal crop worldwide because of its use in the brewing and distilling industry. However, adequate supplies of quality malting barley are threatened by global climate change due to drought in some regions and excess precipitation in others, which facilitates epidemics caused by fungal pathogens. The disease net form net blotch caused by the necrotrophic fungal pathogen Pyrenophora teres f. teres (Ptt) has emerged as a global threat to barley production and diverse populations of Ptt have shown a capacity to overcome deployed genetic resistances. The barley line CI5791 exhibits remarkably effective resistance to diverse Ptt isolates from around the world that maps to two major QTL on chromosomes 3H and 6H. To identify genes involved in this effective resistance, CI5791 seed were γ-irradiated and two mutants, designated CI5791-γ3 and CI5791-γ8, with compromised Ptt resistance were identified from an M2 population. Phenotyping of CI5791-γ3 and -γ8 × Heartland F2 populations showed three resistant to one susceptible segregation ratios and CI5791-γ3 × -γ8 F1 individuals were susceptible, thus these independent mutants are in a single allelic gene. Thirty-four homozygous mutant (susceptible) CI5791-γ3 × Heartland F2 individuals, representing 68 recombinant gametes, were genotyped via PCR genotype by sequencing. The data were used for single marker regression mapping placing the mutation on chromosome 3H within an approximate 75 cM interval encompassing the 3H CI5791 resistance QTL. Sequencing of the mutants and wild-type (WT) CI5791 genomic DNA following exome capture identified independent mutations of the HvWRKY6 transcription factor located on chromosome 3H at ∼50.7 cM, within the genetically delimited region. Post transcriptional gene silencing of HvWRKY6 in barley line CI5791 resulted in Ptt susceptibility, confirming that it functions in NFNB resistance, validating it as the gene underlying the mutant phenotypes. Allele analysis and transcript regulation of HvWRKY6 from resistant and susceptible lines revealed sequence identity and upregulation upon pathogen challenge in all genotypes analyzed, suggesting a conserved transcription factor is involved in the defense against the necrotrophic pathogen. We hypothesize that HvWRKY6 functions as a conserved signaling component of defense mechanisms that restricts Ptt growth in barley.

Research advances in the Pyrenophora teres-barley interaction.

Pyrenophora teres f. teres and P. teres f. maculata are significant pathogens that cause net blotch of barley. An increased number of loci involved in P. teres resistance or susceptibility responses of barley as well as interacting P. teres virulence effector loci have recently been identified through biparental and association mapping studies of both the pathogen and host. Characterization of the resistance/susceptibility loci in the host and the interacting effector loci in the pathogen will provide a path for targeted gene validation for better-informed release of resistant barley cultivars. This review assembles concise consensus maps for all loci published for both the host and pathogen, providing a useful resource for the community to be used in pathogen characterization and barley breeding for resistance to both forms of P. teres.