Diversity of gene expression responses to light quality in barley.

Light quality influence on barley development is poorly understood. We exposed three barley genotypes with either sensitive or insensitive response to two light sources producing different light spectra, fluorescent bulbs, and metal halide lamps, keeping constant light intensity, duration, and temperature. Through RNA-seq, we identified the main genes and pathways involved in the genotypic responses. A first analysis identified genotypic differences in gene expression of development-related genes, including photoreceptors and flowering time genes. Genes from the vernalization pathway of light quality-sensitive genotypes were affected by fluorescent light. In particular, vernalization-related repressors reacted differently: HvVRN2 did not experience relevant changes, whereas HvOS2 expression increased under fluorescent light. To identify the genes primarily related to light quality responses, and avoid the confounding effect of plant developmental stage, genes influenced by development were masked in a second analysis. Quantitative expression levels of PPD-H1, which influenced HvVRN1 and HvFT1, explained genotypic differences in development. Upstream mechanisms (light signaling and circadian clock) were also altered, but no specific genes linking photoreceptors and the photoperiod pathway were identified. The variety of light-quality sensitivities reveals the presence of possible mechanisms of adaptation of winter and facultative barley to latitudinal variation in light quality, which deserves further research.

Sense in sensitivity: difference in the meaning of photoperiod insensitivity between wheat and barley.

The description of long photoperiod sensitivity in wheat and barley is a cause of confusion for researchers working with these crops, usually accustomed to free exchange of physiological and genetic knowledge of such similar crops. Indeed, wheat and barley scientists customarily quote studies of either crop species when researching one of them. Among their numerous similarities, the main gene controlling the long photoperiod sensitivity is the same in both crops (PPD1; PPD-H1 in barley and PPD-D1 in hexaploid wheat). However, the photoperiod responses are different: (i) the main dominant allele inducing shorter time to anthesis is the insensitive allele in wheat (Ppd-D1a) but the sensitive allele in barley (Ppd-H1) (i.e. sensitivity to photoperiod produces opposite effects on time to heading in wheat and barley); (ii) the main 'insensitive' allele in wheat, Ppd-D1a, does confer insensitivity, whilst that of barley reduces the sensitivity but still responds to photoperiod. The different behaviour of PPD1 genes in wheat and barley is put in a common framework based on the similarities and differences of the molecular bases of their mutations, which include polymorphism at gene expression levels, copy number variation, and sequence of coding regions. This common perspective sheds light on a source of confusion for cereal researchers, and prompts us to recommend accounting for the photoperiod sensitivity status of the plant materials when conducting research on genetic control of phenology. Finally, we provide advice to facilitate the management of natural PPD1 diversity in breeding programmes and suggest targets for further modification through gene editing, based on mutual knowledge on the two crops.

Multi-environment genome -wide association mapping of culm morphology traits in barley.

In cereals with hollow internodes, lodging resistance is influenced by morphological characteristics such as internode diameter and culm wall thickness. Despite their relevance, knowledge of the genetic control of these traits and their relationship with lodging is lacking in temperate cereals such as barley. To fill this gap, we developed an image analysis-based protocol to accurately phenotype culm diameters and culm wall thickness across 261 barley accessions. Analysis of culm trait data collected from field trials in seven different environments revealed high heritability values (>50%) for most traits except thickness and stiffness, as well as genotype-by-environment interactions. The collection was structured mainly according to row-type, which had a confounding effect on culm traits as evidenced by phenotypic correlations. Within both row-type subsets, outer diameter and section modulus showed significant negative correlations with lodging (<-0.52 and <-0.45, respectively), but no correlation with plant height, indicating the possibility of improving lodging resistance independent of plant height. Using 50k iSelect SNP genotyping data, we conducted multi-environment genome-wide association studies using mixed model approach across the whole panel and row-type subsets: we identified a total of 192 quantitative trait loci (QTLs) for the studied traits, including subpopulation-specific QTLs and 21 main effect loci for culm diameter and/or section modulus showing effects on lodging without impacting plant height. Providing insights into the genetic architecture of culm morphology in barley and the possible role of candidate genes involved in hormone and cell wall-related pathways, this work supports the potential of loci underpinning culm features to improve lodging resistance and increase barley yield stability under changing environments.

Hybrids Provide More Options for Fine-Tuning Flowering Time Responses of Winter Barley.

Crop adaptation requires matching resource availability to plant development. Tight coordination of the plant cycle with prevailing environmental conditions is crucial to maximizing yield. It is expected that winters in temperate areas will become warmer, so the vernalization requirements of current cultivars can be desynchronized with the environment's vernalizing potential. Therefore, current phenological ideotypes may not be optimum for future climatic conditions. Major genes conferring vernalization sensitivity and phenological responses in barley (Hordeum vulgare L.) are known, but some allelic combinations remain insufficiently evaluated. Furthermore, there is a lack of knowledge about flowering time in a hybrid context. To honor the promise of increased yield potentials, hybrid barley phenology must be studied, and the knowledge deployed in new cultivars. A set of three male and two female barley lines, as well as their six F1 hybrids, were studied in growth chambers, subjected to three vernalization treatments: complete (8 weeks), moderate (4 weeks), and low (2 weeks). Development was recorded up to flowering, and expression of major genes was assayed at key stages. We observed a gradation in responses to vernalization, mostly additive, concentrated in the phase until the initiation of stem elongation, and proportional to the allele constitution and dosage present in VRN-H1. These responses were further modulated by the presence of PPD-H2. The duration of the late reproductive phase presented more dominance toward earliness and was affected by the rich variety of alleles at VRN-H3. Our results provide further opportunities for fine-tuning total and phasal growth duration in hybrid barley, beyond what is currently feasible in inbred cultivars.

Genomic Prediction of Grain Yield in a Barley MAGIC Population Modeling Genotype per Environment Interaction.

Multi-parent Advanced Generation Inter-crosses (MAGIC) lines have mosaic genomes that are generated shuffling the genetic material of the founder parents following pre-defined crossing schemes. In cereal crops, these experimental populations have been extensively used to investigate the genetic bases of several traits and dissect the genetic bases of epistasis. In plants, genomic prediction models are usually fitted using either diverse panels of mostly unrelated accessions or individuals of biparental families and several empirical analyses have been conducted to evaluate the predictive ability of models fitted to these populations using different traits. In this paper, we constructed, genotyped and evaluated a barley MAGIC population of 352 individuals developed with a diverse set of eight founder parents showing contrasting phenotypes for grain yield. We combined phenotypic and genotypic information of this MAGIC population to fit several genomic prediction models which were cross-validated to conduct empirical analyses aimed at examining the predictive ability of these models varying the sizes of training populations. Moreover, several methods to optimize the composition of the training population were also applied to this MAGIC population and cross-validated to estimate the resulting predictive ability. Finally, extensive phenotypic data generated in field trials organized across an ample range of water regimes and climatic conditions in the Mediterranean were used to fit and cross-validate multi-environment genomic prediction models including G×E interaction, using both genomic best linear unbiased prediction and reproducing kernel Hilbert space along with a non-linear Gaussian Kernel. Overall, our empirical analyses showed that genomic prediction models trained with a limited number of MAGIC lines can be used to predict grain yield with values of predictive ability that vary from 0.25 to 0.60 and that beyond QTL mapping and analysis of epistatic effects, MAGIC population might be used to successfully fit genomic prediction models. We concluded that for grain yield, the single-environment genomic prediction models examined in this study are equivalent in terms of predictive ability while, in general, multi-environment models that explicitly split marker effects in main and environmental-specific effects outperform simpler multi-environment models.

Major flowering time genes of barley: allelic diversity, effects, and comparison with wheat.

KEY MESSAGE: This review summarizes the allelic series, effects, interactions between genes and with the environment, for the major flowering time genes that drive phenological adaptation of barley. The optimization of phenology is a major goal of plant breeding addressing the production of high-yielding varieties adapted to changing climatic conditions. Flowering time in cereals is regulated by genetic networks that respond predominately to day length and temperature. Allelic diversity at these genes is at the basis of barley wide adaptation. Detailed knowledge of their effects, and genetic and environmental interactions will facilitate plant breeders manipulating flowering time in cereal germplasm enhancement, by exploiting appropriate gene combinations. This review describes a catalogue of alleles found in QTL studies by barley geneticists, corresponding to the genetic diversity at major flowering time genes, the main drivers of barley phenological adaptation: VRN-H1 (HvBM5A), VRN-H2 (HvZCCTa-c), VRN-H3 (HvFT1), PPD-H1 (HvPRR37), PPD-H2 (HvFT3), and eam6/eps2 (HvCEN). For each gene, allelic series, size and direction of QTL effects, interactions between genes and with the environment are presented. Pleiotropic effects on agronomically important traits such as grain yield are also discussed. The review includes brief comments on additional genes with large effects on phenology that became relevant in modern barley breeding. The parallelisms between flowering time allelic variation between the two most cultivated Triticeae species (barley and wheat) are also outlined. This work is mostly based on previously published data, although we added some new data and hypothesis supported by a number of studies. This review shows the wide variety of allelic effects that provide enormous plasticity in barley flowering behavior, which opens new avenues to breeders for fine-tuning phenology of the barley crop.

Responses of Barley to High Ambient Temperature Are Modulated by Vernalization.

Ambient temperatures are increasing due to climate change. Cereal crops development and production will be affected consequently. Flowering time is a key factor for adaptation of small grain cereals and, therefore, exploring developmental responses of barley to rising temperatures is required. In this work, we studied phasic growth, and inflorescence traits related to yield, in eight near isogenic lines of barley (Hordeum vulgare L.) differing at the VRN-H1, VRN-H2 and PPD-H1 genes, representing different growth habits. The lines were grown in contrasting vernalization treatments, under two temperature regimes (18 and 25°C), in long days. Lines with recessive ppd-H1 presented delayed development compared to lines with the sensitive PPD-H1 allele, across the two growth phases considered. High temperature delayed flowering in all unvernalized plants, and in vernalized spring barleys carrying the insensitive ppd-H1 allele, whilst it accelerated flowering in spring barleys with the sensitive PPD-H1 allele. This finding evidenced an interaction between PPD-H1, temperature and vernalization. At the high temperature, PPD-H1 lines in spring backgrounds (VRN-H1-7) yielded more, whereas lines with ppd-H1 were best in vrn-H1 background. Our study revealed new information that will support breeding high-yielding cultivars with specific combinations of major adaptation genes tailored to future climatic conditions.

Genetic diversity in developmental responses to light spectral quality in barley (Hordeum vulgare L.).

BACKGROUND: Plants use light wavelength, intensity, direction and duration to predict imminent seasonal changes and to determine when to initiate physiological and developmental processes. Among them, crop responses to light are not fully understood. Here, we study how light quality affects barley development, using two broad-spectrum light sources, metal halide (M) and fluorescent (F) lamps. Eleven varieties with known allelic variants for the major flowering time genes were evaluated under controlled conditions (long days, same light intensity). Two experiments were carried out with fully-vernalized plants: 1) control treatments (M, F); 2) shifting chambers 10 days after the start of the experiment (MF, FM). RESULTS: In general, varieties developed faster under longer exposure to M conditions. The greatest differences were due to a delay promoted by F light bulbs, especially in the time to first node appearance and until the onset of stem elongation. Yield related-traits as the number of seeds were also affected by the conditions experienced. However, not each variety responded equally, and they could be classified in insensitive and sensitive to light quality. Expression levels of flowering time genes HvVRN1, HvFT1 and PPD-H1 were high in M, while HvFT3 and HvVRN2 were higher under F conditions. The expression under shift treatments revealed also a high correlation between HvVRN1 and PPD-H1 transcript levels. CONCLUSIONS: The characterization of light quality effects has highlighted the important influence of the spectrum on early developmental stages, affecting the moment of onset of stem elongation, and further consequences on the morphology of the plant and yield components. We suggest that light spectra control the vernalization and photoperiod genes probably through the regulation of upstream elements of signalling pathways. The players behind the different responses to light spectra found deserve further research, which could help to optimize breeding strategies.

Rapid On-Site Phenotyping via Field Fluorimeter Detects Differences in Photosynthetic Performance in a Hybrid-Parent Barley Germplasm Set.

Crop productivity can be expressed as the product of the amount of radiation intercepted, radiation use efficiency and harvest index. Genetic variation for components of radiation use efficiency has rarely been explored due to the lack of appropriate equipment to determine parameters at the scale needed in plant breeding. On the other hand, responses of the photosynthetic apparatus to environmental conditions have not been extensively investigated under field conditions, due to the challenges posed by the fluctuating environmental conditions. This study applies a rapid, low-cost, and reliable high-throughput phenotyping tool to explore genotypic variation for photosynthetic performance of a set of hybrid barleys and their parents under mild water-stress and unstressed field conditions. We found differences among the genotypic sets that are relevant for plant breeders and geneticists. Hybrids showed lower leaf temperature differential and higher non-photochemical quenching, resembling closer the male parents. The combination of traits detected in hybrids seems favorable, and could indicate improved photoprotection and better fitness under stress conditions. Additionally, we proved the potential of a low-cost, field-based phenotyping equipment to be used routinely in barley breeding programs for early screening for stress tolerance.

Perspectives on Low Temperature Tolerance and Vernalization Sensitivity in Barley: Prospects for Facultative Growth Habit.

One option to achieving greater resiliency for barley production in the face of climate change is to explore the potential of winter and facultative growth habits: for both types, low temperature tolerance (LTT) and vernalization sensitivity are key traits. Sensitivity to short-day photoperiod is a desirable attribute for facultative types. In order to broaden our understanding of the genetics of these phenotypes, we mapped quantitative trait loci (QTLs) and identified candidate genes using a genome-wide association studies (GWAS) panel composed of 882 barley accessions that was genotyped with the Illumina 9K single-nucleotide polymorphism (SNP) chip. Fifteen loci including 5 known and 10 novel QTL/genes were identified for LTT-assessed as winter survival in 10 field tests and mapped using a GWAS meta-analysis. FR-H1, FR-H2, and FR-H3 were major drivers of LTT, and candidate genes were identified for FR-H3. The principal determinants of vernalization sensitivity were VRN-H1, VRN-H2, and PPD-H1. VRN-H2 deletions conferred insensitive or intermediate sensitivity to vernalization. A subset of accessions with maximum LTT were identified as a resource for allele mining and further characterization. Facultative types comprised a small portion of the GWAS panel but may be useful for developing germplasm with this growth habit.

Harnessing Novel Diversity From Landraces to Improve an Elite Barley Variety.

The Spanish Barley Core Collection (SBCC) is a source of genetic variability of potential interest for breeding, particularly for adaptation to Mediterranean environments. Two backcross populations (BC2F5) were developed using the elite cultivar Cierzo as the recurrent parent. The donor parents, namely SBCC042 and SBCC073, were selected from the SBCC lines due to their outstanding yield in drought environments. Flowering time, yield and drought-related traits were evaluated in two field trials in Zaragoza (Spain) during the 2014-15 and 2015-16 seasons and validated in the 2017-18 season. Two hundred sixty-four lines of each population were genotyped with the Barley Illumina iSelect 50k SNP chip. Genetic maps for each population were generated. The map for SBCC042 × Cierzo contains 12,893 SNPs distributed in 9 linkage groups. The map for SBCC073 × Cierzo includes 12,026 SNPs in 7 linkage groups. Both populations shared two QTL hotspots. There are QTLs for flowering time, thousand-kernel weight (TKW), and hectoliter weight on a segment of 23 Mb at ~515 Mb on chromosome 1H, which encompasses the HvFT3 gene. In both populations, flowering was accelerated by the landrace allele, which also increased the TKW. In the same region, better soil coverage was contributed by SBCC042 but coincident with a lower hectoliter weight. The second large hotspot was on chromosome 6H and contained QTLs with wide intervals for grain yield, plant height and TKW. Landrace alleles contributed to increased plant height and TKW and reduced grain yield. Only SBCC042 contributed favorable alleles for "green area," with three significant QTLs that increased ground coverage after winter, which might be exploited as an adaptive trait of this landrace. Some genes of interest found in or very close to the peaks of the QTLs are highlighted. Strategies to deploy the QTLs found for breeding and pre-breeding are proposed.

Fine-tuning of the flowering time control in winter barley: the importance of HvOS2 and HvVRN2 in non-inductive conditions.

BACKGROUND: In winter barley plants, vernalization and photoperiod cues have to be integrated to promote flowering. Plant development and expression of different flowering promoter (HvVRN1, HvCO2, PPD-H1, HvFT1, HvFT3) and repressor (HvVRN2, HvCO9 and HvOS2) genes were evaluated in two winter barley varieties under: (1) natural increasing photoperiod, without vernalization, and (2) under short day conditions in three insufficient vernalization treatments. These challenging conditions were chosen to capture non-optimal and natural responses, representative of those experienced in the Mediterranean area. RESULTS: In absence of vernalization and under increasing photoperiods, HvVRN2 expression increased with day-length, mainly between 12 and 13 h photoperiods in our latitudes. The flowering promoter gene in short days, HvFT3, was only expressed after receiving induction of cold or plant age, which was associated with low transcript levels of HvVRN2 and HvOS2. Under the sub-optimal conditions here described, great differences in development were found between the two winter barley varieties used in the study. Delayed development in 'Barberousse' was associated with increased expression levels of HvOS2. Novel variation for HvCO9 and HvOS2 is reported and might explain such differences. CONCLUSIONS: The balance between the expression of flowering promoters and repressor genes regulates the promotion towards flowering or the maintenance of the vegetative state. HvOS2, an ortholog of FLC, appears as a strong candidate to mediate in the vernalization response of barley. Natural variation found would help to exploit the plasticity in development to obtain better-adapted varieties for current and future climate conditions.

Genetic association with high-resolution climate data reveals selection footprints in the genomes of barley landraces across the Iberian Peninsula.

Landraces are local populations of crop plants adapted to a particular environment. Extant landraces are surviving genetic archives, keeping signatures of the selection processes experienced by them until settling in their current niches. This study intends to establish relationships between genetic diversity of barley (Hordeum vulgare L.) landraces collected in Spain and the climate of their collection sites. A high-resolution climatic data set (5 × 5 km spatial, 1-day temporal grid) was computed from over 2,000 temperature and 7,000 precipitation stations across peninsular Spain. This data set, spanning the period 1981-2010, was used to derive agroclimatic variables meaningful for cereal production at the collection sites of 135 barley landraces. Variables summarize temperature, precipitation, evapotranspiration, potential vernalization and frost probability at different times of the year and time scales (season and month). SNP genotyping of the landraces was carried out combining Illumina Infinium assays and genotyping-by-sequencing, yielding 9,920 biallelic markers (7,479 with position on the barley reference genome). The association of these SNPs with agroclimatic variables was analysed at two levels of genetic diversity, with and without taking into account population structure. The whole data sets and analysis pipelines are documented and available at https://eead-csic-compbio.github.io/barley-agroclimatic-association. We found differential adaptation of the germplasm groups identified to be dominated by reactions to cold temperature and late-season frost occurrence, as well as to water availability. Several significant associations pointing at specific adaptations to agroclimatic features related to temperature and water availability were observed, and candidate genes underlying some of the main regions are proposed.

Large Differences in Gene Expression Responses to Drought and Heat Stress between Elite Barley Cultivar Scarlett and a Spanish Landrace.

Drought causes important losses in crop production every season. Improvement for drought tolerance could take advantage of the diversity held in germplasm collections, much of which has not been incorporated yet into modern breeding. Spanish landraces constitute a promising resource for barley breeding, as they were widely grown until last century and still show good yielding ability under stress. Here, we study the transcriptome expression landscape in two genotypes, an outstanding Spanish landrace-derived inbred line (SBCC073) and a modern cultivar (Scarlett). Gene expression of adult plants after prolonged stresses, either drought or drought combined with heat, was monitored. Transcriptome of mature leaves presented little changes under severe drought, whereas abundant gene expression changes were observed under combined mild drought and heat. Developing inflorescences of SBCC073 exhibited mostly unaltered gene expression, whereas numerous changes were found in the same tissues for Scarlett. Genotypic differences in physiological traits and gene expression patterns confirmed the different behavior of landrace SBCC073 and cultivar Scarlett under abiotic stress, suggesting that they responded to stress following different strategies. A comparison with related studies in barley, addressing gene expression responses to drought, revealed common biological processes, but moderate agreement regarding individual differentially expressed transcripts. Special emphasis was put in the search of co-expressed genes and underlying common regulatory motifs. Overall, 11 transcription factors were identified, and one of them matched cis-regulatory motifs discovered upstream of co-expressed genes involved in those responses.

Analysis of Plant Pan-Genomes and Transcriptomes with GET_HOMOLOGUES-EST, a Clustering Solution for Sequences of the Same Species.

The pan-genome of a species is defined as the union of all the genes and non-coding sequences found in all its individuals. However, constructing a pan-genome for plants with large genomes is daunting both in sequencing cost and the scale of the required computational analysis. A more affordable alternative is to focus on the genic repertoire by using transcriptomic data. Here, the software GET_HOMOLOGUES-EST was benchmarked with genomic and RNA-seq data of 19 Arabidopsis thaliana ecotypes and then applied to the analysis of transcripts from 16 Hordeum vulgare genotypes. The goal was to sample their pan-genomes and classify sequences as core, if detected in all accessions, or accessory, when absent in some of them. The resulting sequence clusters were used to simulate pan-genome growth, and to compile Average Nucleotide Identity matrices that summarize intra-species variation. Although transcripts were found to under-estimate pan-genome size by at least 10%, we concluded that clusters of expressed sequences can recapitulate phylogeny and reproduce two properties observed in A. thaliana gene models: accessory loci show lower expression and higher non-synonymous substitution rates than core genes. Finally, accessory sequences were observed to preferentially encode transposon components in both species, plus disease resistance genes in cultivated barleys, and a variety of protein domains from other families that appear frequently associated with presence/absence variation in the literature. These results demonstrate that pan-genome analyses are useful to explore germplasm diversity.

A Cluster of Nucleotide-Binding Site-Leucine-Rich Repeat Genes Resides in a Barley Powdery Mildew Resistance Quantitative Trait Loci on 7HL.

Powdery mildew causes severe yield losses in barley production worldwide. Although many resistance genes have been described, only a few have already been cloned. A strong QTL (quantitative trait locus) conferring resistance to a wide array of powdery mildew isolates was identified in a Spanish barley landrace on the long arm of chromosome 7H. Previous studies narrowed down the QTL position, but were unable to identify candidate genes or physically locate the resistance. In this study, the exome of three recombinant lines from a high-resolution mapping population was sequenced and analyzed, narrowing the position of the resistance down to a single physical contig. Closer inspection of the region revealed a cluster of closely related NBS-LRR (nucleotide-binding site-leucine-rich repeat containing protein) genes. Large differences were found between the resistant lines and the reference genome of cultivar Morex, in the form of PAV (presence-absence variation) in the composition of the NBS-LRR cluster. Finally, a template-guided assembly was performed and subsequent expression analysis revealed that one of the new assembled candidate genes is transcribed. In summary, the results suggest that NBS-LRR genes, absent from the reference and the susceptible genotypes, could be functional and responsible for the powdery mildew resistance. The procedure followed is an example of the use of NGS (next-generation sequencing) tools to tackle the challenges of gene cloning when the target gene is absent from the reference genome.

HvFT1 polymorphism and effect-survey of barley germplasm and expression analysis.

Flowering time in plants is a tightly regulated process. In barley (Hordeum vulgare L.), HvFT1, ortholog of FLOWERING LOCUS T, is the main integrator of the photoperiod and vernalization signals leading to the transition from vegetative to reproductive state of the plant. This gene presents sequence polymorphisms affecting flowering time in the first intron and in the promoter. Recently, copy number variation (CNV) has been described for this gene. An allele with more than one copy was linked to higher gene expression, earlier flowering, and an overriding effect of the vernalization mechanism. This study aims at (1) surveying the distribution of HvFT1 polymorphisms across barley germplasm and (2) assessing gene expression and phenotypic effects of HvFT1 alleles. We analyzed HvFT1 CNV in 109 winter, spring, and facultative barley lines. There was more than one copy of the gene (2-5) only in spring or facultative barleys without a functional vernalization VrnH2 allele. CNV was investigated in several regions inside and around HvFT1. Two models of the gene were found: one with the same number of promoters and transcribed regions, and another with one promoter and variable number of transcribed regions. This last model was found in Nordic barleys only. Analysis of HvFT1 expression showed that association between known polymorphisms at the HvFT1 locus and the expression of the gene was highly dependent on the genetic background. Under long day conditions the earliest flowering lines carried a sensitive PpdH1 allele. Among spring cultivars with different number of copies, no clear relation was found between CNV, gene expression and flowering time. This was confirmed in a set of doubled haploid lines of a population segregating for HvFT1 CNV. Earlier flowering in the presence of several copies of HvFT1 was only seen in cultivar Tammi, which carries one promoter, suggesting a relation of gene structure with its regulation. HvCEN also affected to a large extent flowering time.

Fine mapping of the Rrs1 resistance locus against scald in two large populations derived from Spanish barley landraces.

KEY MESSAGE: In two Spanish barley landraces with outstanding resistance to scald, the Rrs1 Rh4 locus was fine mapped including all known markers used in previous studies and closely linked markers were developed. Scald, caused by Rhynchosporium commune, is one of the most prevalent barley diseases worldwide. A search for new resistance sources revealed that Spanish landrace-derived lines SBCC145 and SBCC154 showed outstanding resistance to scald. They were crossed to susceptible cultivar Beatrix to create large DH-mapping populations of 522 and 416 DH lines that were scored for disease resistance in the greenhouse using two R. commune isolates. To ascertain the pattern of resistance, parents and reference barley lines with known scald resistance were phenotyped with a panel of differential R. commune isolates. Subpopulations were genotyped with the Illumina GoldenGate 1,536 SNP Assay and a large QTL in the centromeric region of chromosome 3H, known to harbour several scald resistance genes and/or alleles, was found in both populations. Five SNP markers closest to the QTL were converted into CAPS markers. These CAPS markers, together with informative SSR markers used in other scald studies, confirmed the presence of the Rrs1 locus. The panel of differential scald isolates indicated that the allele carried by both donors was Rrs1 Rh4 . The genetic distance between Rrs1 and its flanking markers was 1.2 cM (11_0010) proximally and 0.9 cM (11_0823) distally, which corresponds to a distance of just below 9 Mbp. The number and nature of scald resistance genes on chromosome 3H are discussed. The effective Rrs1 allele found and the closely linked markers developed are already useful tools for molecular breeding programs and provide a good step towards the identification of candidate genes.

Towards positional isolation of three quantitative trait loci conferring resistance to powdery mildew in two Spanish barley landraces.

Three quantitative trait loci (QTL) conferring broad spectrum resistance to powdery mildew, caused by the fungus Blumeria graminis f. sp. hordei, were previously identified on chromosomes 7HS, 7HL and 6HL in the Spanish barley landrace-derived lines SBCC097 and SBCC145. In the present work, a genome-wide putative linear gene index of barley (Genome Zipper) and the first draft of the physical, genetic and functional sequence of the barley genome were used to go one step further in the shortening and explicit demarcation on the barley genome of these regions conferring resistance to powdery mildew as well as in the identification of candidate genes. First, a comparative analysis of the target regions to the barley Genome Zippers of chromosomes 7H and 6H allowed the development of 25 new gene-based molecular markers, which slightly better delimit the QTL intervals. These new markers provided the framework for anchoring of genetic and physical maps, figuring out the outline of the barley genome at the target regions in SBCC097 and SBCC145. The outermost flanking markers of QTLs on 7HS, 7HL and 6HL defined a physical area of 4 Mb, 3.7 Mb and 3.2 Mb, respectively. In total, 21, 10 and 16 genes on 7HS, 7HL and 6HL, respectively, could be interpreted as potential candidates to explain the resistance to powdery mildew, as they encode proteins of related functions with respect to the known pathogen defense-related processes. The majority of these were annotated as belonging to the NBS-LRR class or protein kinase family.