M. P. Gracia

The Spanish barley core collection

Spanish barleys constitute a germplasm group of particular interest for breeding purposes, as Spain has been proposed as a possible centre of origin of the crop. The Spanish National Germplasm Bank (Banco Nacional de Germoplasma, BNG), holds a collection of about 2000 barley accessions, mostly landraces collected in Spain prior to extensive introduction of modern varieties. The objective of this work is to create a core collection of barleys representative of old barley genotypes grown in Spain. The core collection will be constituted by three groups of germplasm: successful old varieties (15); entries in common with previously existing barley core collections (15); and 2-row (8) and 6-row (122) entries from the BNG, for a total of 160 entries. Entries were allocated by stratified sampling in agro-ecological uniform zones of barley cultivation in Spain. Classification of agro-ecological regions for barley was based on historical yield records for Spanish provinces. The number of entries for each region was determined in proportion to the logarithm of historical barley acreage. Final choice of accessions within provinces tried to maximize the diversity and avoid duplications by looking at passport data, and to agronomic evaluation data available for a group of about 900 accessions.

Patterns of genetic and eco-geographical diversity in Spanish barleys

The pool of Western Mediterranean landraces has been under-utilised for barley breeding so far. The objectives of this study were to assess genetic diversity in a core collection of inbred lines derived from Spanish barley landraces to establish its relationship to barleys from other origins, and to correlate the distribution of diversity with geographical and climatic factors. To this end, 64 SSR were used to evaluate the polymorphism among 225 barley (Hordeum vulgare ssp. vulgare) genotypes, comprising two-row and six-row types. These included 159 landraces from the Spanish barley core collection (SBCC) plus 66 cultivars, mainly from European countries, as a reference set. Out of the 669 alleles generated, a large proportion of them were unique to the six-row Spanish barleys. An analysis of molecular variance revealed a clear genetic divergence between the six-row Spanish barleys and the reference cultivars, whereas this was not evident for the two-row barleys. A model-based clustering analysis identified an underlying population structure, consisting of four main populations for the whole genotype set, and suggested further possible subdivision within two of these populations. Most of the six-row Spanish landraces clustered into two groups that corresponded to geographic regions with contrasting environmental conditions. The existence of wide genetic diversity in Spanish germplasm, possibly related to adaptation to a broad range of environmental conditions, and its divergence from current European cultivars confirm its potential as a new resource for barley breeders, and make the SBCC a valuable tool for the study of adaptation in barley.

Heading date QTL in a spring × winter barley cross evaluated in Mediterranean environments

Heading date is a key trait for the adaptation of barley to Mediterranean environments. We studied the genetic control of flowering time under Northern Spanish (Mediterranean) conditions using a new population derived from the spring/winter cross Beka/Mogador. A set of 120 doubled haploid lines was evaluated in the field, and under controlled temperature and photoperiod conditions. Genotyping was carried out with 215 markers (RFLP, STS, RAPD, AFLP, SSR), including markers for vernalization candidate genes, HvBM5 (Vrn-H1), HvZCCT (Vrn-H2), and HvT SNP22 (Ppd-H1). Four major QTL, and the interactions between them, accounted for most of the variation in both field (71–92%) and greenhouse trials (55–86%). These were coincident with the location of the major genes for response to vernalization and short photoperiod (Ppd-H2 on chromosome 1H). A major QTL, near the centromere of chromosome 2H was the most important under autumn sowing conditions. Although it is detected under all conditions, its action seems not independent from environmental cues. An epistatic interaction involving the two vernalization genes was detected when the plants were grown without vernalization and under long photoperiod. The simultaneous presence of the winter Mogador allele at the two loci produced a marked delay in heading date, beyond a mere additive effect. This interaction, combined with the effect of the gene responsive to short photoperiod, Ppd-H2, was found responsible of the phenomenon known as short-day vernalization, present in some of the lines of the population.

Field responses of grain sorghum to a salinity gradient

Abstract Grain sorghum is a potential crop for moderately saline areas, having been identified as fairly tolerant to salinity, and shown to contain intraspecific variability for that trait. The aim of this work was to describe the responses of grain sorghum to saline irrigation, assess the responses of a set of genotypes to salinity, and to analyze the relationships between several agronomic and physiological traits and salinity tolerance. In an experiment during three years, eleven public inbred lines and one cultivar were exposed to a salinity gradient (NaCl and CaCl2, 1:1 w/w) created with a triple line source sprinkler system. The traits most affected by salinity were grain yield, number of grains per head, shoot dry weight (both grain and stover), harvest index, and leaf chloride, sodium, calcium, and potassium concentrations. Plant height, head length, and head number per plot were moderately affected by salinity, whereas flowering time, and total number of leaves per plant were unaffected. Two sets of three genotypes were identified with consistently contrasting responses to salinity across the three years. The differences in tolerance between these two groups were not associated with differences in total shoot biomass, but rather with different patterns of biomass partitioning under the most saline conditions. There were significant differences between the tolerant and susceptible genotypes in leaf chloride and potassium concentrations. The possible implications of the latter in the determination of the contrasting genotypic responses to salinity are discussed.

CHARACTERIZATION AND GENETIC CONTROL OF GERMINATION-EMERGENCE RESPONSES OF GRAIN SORGHUM TO SALINITY

SummaryWhen grain sorghum is grown in saline soils, one cause of low yield is poor crop establishment. The objectives of this study were to assess the response of grain sorghum to salinity in the germination-emergence stages, study the inheritance of salt tolerance at this stage, and determine the relative contribution to final emergence of salt effects during imbibition, and after onset of germination. Twelve inbred lines and 18 F1 hybrids, resulting from an incomplete 6×6 factorial mating design, were tested for germination and emergence in folded paper at 10 salt concentrations, from 1.8 to 36 dSm-1. The mean EC50 (the electrical conductivity at which the variable score declines by 50%) for emerged seedlings production was 21.2 dSm-1. Large genotypic differences were observed for salt tolerance at germination and emergence stages, which were not related to the viability of seeds, and poorly related to seed weight (considered as an estimate of intrinsic seed vigor). In the hybrids, these differences were due to SCA and female GCA for emergence, and female GCA for germination, though the male GCA was also significant for both characters. Line per se performance was significantly correlated to individual GCA estimates for emergence, but not for germination. Heterosis was only detected in three crosses for final emergence and in one cross for germination. The genetic differences in final emergence were mainly due to effects occurring after the onset of germination rather than a consequence of effects during imbibition.

Resistance to powdery mildew in Spanish barley landraces is controlled by different sets of quantitative trait loci

Twenty-two landrace-derived inbred lines from the Spanish Barley Core Collection (SBCC) were found to display high levels of resistance to a panel of 27 isolates of the fungus Blumeria graminis that exhibit a wide variety of virulences. Among these lines, SBCC145 showed high overall resistance and a distinctive spectrum of resistance compared with the other lines. Against this background, the main goal of the present work was to investigate the genetic basis underlying such resistance using a doubled haploid population derived from a cross between SBCC145 and the elite spring cultivar Beatrix. The population was genotyped with the 1,536-SNP Illumina GoldenGate Oligonucleotide Pool Assay (Barley OPA-1 or BOPA1 for short), whereas phenotypic analysis was performed using two B. graminis isolates. A major quantitative trait locus (QTL) for resistance to both isolates was identified on the long arm of chromosome 6H (6HL) and accounted for ca. 60% of the phenotypic variance. Depending on the B. graminis isolate tested, three other minor QTLs were detected on chromosomes 2H and 7H, which explained less than 5% of the phenotypic variation each. In all cases, the alleles for resistance derived from the Spanish parent SBCC145. The position, the magnitude of the effect observed and the proportion of phenotypic variation accounted for by the QTL on 6HL suggest this is a newly identified locus for broad-based resistance to powdery mildew.

Identification of quantitative trait loci for resistance to powdery mildew in a Spanish barley landrace

The Spanish landrace-derived inbred line SBCC97, together with other lines from the Spanish Barley Core Collection, displays high resistance to powdery mildew, caused by the fungus Blumeria graminis f. sp. hordei. The objective of this study was to map quantitative trait loci (QTLs) for resistance to powdery mildew in a recombinant inbred line population derived from a cross between SBCC97 and the susceptible cultivar ‘Plaisant’. Phenotypic analysis was performed using four B. graminis isolates, and genetic maps were constructed with mainly simple sequence repeat (SSR) markers, following a sequential genotyping strategy. Two major QTLs with large effects were identified on chromosome 7H, and they accounted for up to 45% of the total phenotypic variance. The alleles for resistance at each QTL were contributed by the Spanish parent SBCC97. One locus was mapped to the short arm of chromosome 7HS, and was flanked by the resistance gene analogue (RGA) marker S9202 and the SSR GBM1060. This corresponded to the same chromosomal region in which a major race-specific resistance gene from Hordeum vulgare ssp. spontaneum, designated as mlt, had been identified previously. The second QTL was linked tightly to marker EBmac0755, and it shared its chromosomal location with the qualitative resistance gene Mlf, which has only been described previously in the wild ancestor H. spontaneum. This is the first report of these two QTLs occurring together in cultivated barley, and it paves the way for their use in barley breeding programs that are designed to transfer resistance alleles into elite cultivars.

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.

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.

Resequencing theVrs1 gene in Spanish barley landraces revealed reversion of six-rowed to two-rowed spike

Six-rowed spike 1 (Vrs1) is a gene of major importance for barley breeding and germplasm management as it is the main gene determining spike row-type (2-rowed vs. 6-rowed). This is a widely used DUS trait, and has been often associated to phenotypic traits beyond spike type. Comprehensive re-sequencing Vrs1 revealed three two-rowed alleles (Vrs1.b2; Vrs1.b3; Vrs1.t1) and four six-rowed (vrs1.a1; vrs1.a2; vrs1.a3; vrs1.a4) in the natural population. However, the current knowledge about Vrs1 alleles and its distribution among Spanish barley subpopulations is still underexploited. We analyzed the gene in a panel of 215 genotypes, made of Spanish landraces and European cultivars. Among 143 six-rowed accessions, 57 had the vrs1.a1 allele, 83 were vrs1.a2, and three showed the vrs1.a3 allele. Vrs1.b3 was found in most two-rowed accessions, and a new allele was observed in 7 out of 50 two-rowed Spanish landraces. This allele, named Vrs1.b5, contains a ‘T’ insertion in exon 2, originally proposed as the causal mutation giving rise to the six-row vrs1.a2 allele, but has an additional upstream deletion that results in the change of 15 amino acids and a potentially functional protein. We conclude that eight Vrs1 alleles (Vrs1.b2, Vrs1.b3, Vrs1.b5, Vrs1.t1, vrs1.a1, vrs1.a2, vrs1.a3, vrs1.a4) discriminate two and six-rowed barleys. The markers described will be useful for DUS identification, plant breeders, and other crop scientists.