Ernesto Igartua

Genetic diversity of Prunus rootstocks analyzed by RAPD markers

We have used RAPD markers to characterize Prunus rootstocks from different species, both commercial, and selected clones from the breeding program at Aula Dei Experimental Station (Zaragoza, Spain). Molecular markers were used to study the genetic variation among different species, and within species. Forty one genotypes were used in this study. They included P. amygdalo-persica, and P. persica × P. davidiana hybrids; P. cerasifera, P. domestica, and P. insititia clones, and other diverse interspecific hybrids, which were divided in three groups according to postulated taxonomic classification. Diversity patterns obtained from 80 RAPD primers were evaluated in a representative subset of genotypes. This screening helped to identify 7 RAPD primers that were selected to produce a combined classification of the whole set of rootstock clones. This analysis successfully clustered rootstocks according to the classification scheme widely used to characterize Prunus clones, mainly based on morphological descriptors. Further than that, it supported the alleged origin of some interspecific materials, and confirmed a case of possible misclassification (‘Myrobalan 29 C’). A more thorough diversity analysis was conducted within each group of materials, using larger sets of primers (12–14). After this analysis, disjointed clusters were formed for P. amygdalo-persica and P. persica × P. davidiana hybrids in one group, and for Myrobalan (P. cerasifera) and Marianna (P. cerasifera × P. munsoniana) plums in another group. P. insititia and P. domestica clones, however, formed a jumbled cluster, possibly due to genetic interchange among them during their domestication and breeding history.

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.

Further evidence supporting Morocco as a centre of origin of barley

Abstract Thirty-five populations of H. spontaneum from nine countries, encompassing almost all the known range of distribution of the species, Afghanistan, Crete (Greece), Cyprus, Iran, Iraq, Israel, Libya, Morocco and Turkey, were studied utilizing RFLP markers (21 probes with three restriction enzymes) distributed across all seven barley chromosomes in an attempt to unveil the genetic dissimilarities existing among them. UPGMA clustering, based on the Nei and Li (1979) similarity coefficient, produced a dendrogram where three clusters could be defined: two with a clear geographical distinction (Morocco and Cyprus) and another one grouping all the Asian/Middle Eastern populations, except for an accession from Iran that clustered separately. These results confirm our previous work and suggest that barley domestication could also have taken place outside the Fertile Crescent, particularly in Morocco.

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.

Expression analysis of vernalization and day-length response genes in barley (Hordeum vulgare L.) indicates that VRNH2 is a repressor of PPDH2 (HvFT3) under long days

The response to vernalization and the expression of genes associated with responses to vernalization (VRNH1, VRNH2, and VRNH3) and photoperiod (PPDH1 and PPDH2) were analysed in four barley (Hordeum vulgare L.) lines: ‘Alexis’ (spring), ‘Plaisant’ (winter), SBCC058, and SBCC106 (Spanish inbred lines), grown under conditions of vernalization and short days (VSD) or no vernalization and long days (NVLD). The four genotypes differ in VRNH1. Their growth habits and responses to vernalization correlated with the level of expression of VRNH1 and the length of intron 1. ‘Alexis’ and ‘Plaisant’ behaved as expected. SBCC058 and SBCC106 showed an intermediate growth habit and flowered relatively late in the absence of vernalization. VRNH1 expression was induced by cold for all genotypes. Under VSD, VRNH1 expression was detected in the SBCC genotypes later than in ‘Alexis’ but earlier than in ‘Plaisant’. VRNH2 was repressed under short days while VRNH1 expression increased in parallel. VRNH3 was detected only in ‘Alexis’ under NVLD, whereas it was not expressed in plants with the active allele of VRNH2 (SBCC058 and ‘Plaisant’). Under VSD, PPDH2 was expressed in ‘Alexis’, SBCC058, and SBCC106, but it was only expressed weakly in ‘Alexis’ under NVLD. Further analysis of PPDH2 expression in two barley doubled haploid populations revealed that, under long days, HvFT3 and VRNH2 expression levels were related inversely. The timing of VRNH2 expression under a long photoperiod suggests that this gene might be involved in repression of PPDH2 and, indirectly, in the regulation of flowering time through an interaction with the day-length pathway.

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.

Prognosis of iron chlorosis from the mineral composition of flowers in peach

Summary A statistical approach has been applied to study the nutrient composition of flowers in a peach orchard with 50 peach trees suffering from different degrees of Fe deficiency. Flowers from each individual tree were analysed in five crop seasons. The existence of a consistent nutrient balance over years in peach flowers, with nutrient concentrations depending to some extent on the levels of other nutrients, was revealed by a principal component-type analysis. The analysis produced a dominant first principal component, which explained 21% of the total variance. The second component explained 13% of the variance, with further components explaining less than 9%. A regression analysis revealed that the first principal component was closely related to chlorophyll (Chl) concentration determined in leaves 120 d after full bloom, as it explained 59% of average Chl concentration variance (between 40 and 52% in single year analysis), which is equivalent to a correlation coefficient of 0.77. A multiple regression model using the flower concentrations of N, P, K, Ca, Mg, Fe, Mn, Cu and Zn explained from 50 to 62% of the Chl variation among trees in the different seasons. Flower Fe was significantly correlated with Chl some years, whereas in other years the correlation was weak. Regression models including K and Zn explained 27 to 29% of leaf Chl concentration variance. This coefficient of determination is equivalent to a correlation coefficient of -0.52. This relationship was quite constant across years. We propose that the K/Zn ratio in flowers could be used, along with the flower Fe concentration, for the prognosis of iron chlorosis in peach trees.

Adaptation of barley to mild winters: A role for PPDH2

BackgroundUnderstanding the adaptation of cereals to environmental conditions is one of the key areas in which plant science can contribute to tackling challenges presented by climate change. Temperature and day length are the main environmental regulators of flowering and drivers of adaptation in temperate cereals. The major genes that control flowering time in barley in response to environmental cues are VRNH1, VRNH2, VRNH3, PPDH1, and PPDH2 (candidate gene HvFT3). These genes from the vernalization and photoperiod pathways show complex interactions to promote flowering that are still not understood fully. In particular, PPDH2 function is assumed to be limited to the ability of a short photoperiod to promote flowering. Evidence from the fields of biodiversity, ecogeography, agronomy, and molecular genetics was combined to obtain a more complete overview of the potential role of PPDH2 in environmental adaptation in barley.ResultsThe dominant PPDH2 allele is represented widely in spring barley cultivars but is found only occasionally in modern winter cultivars that have strong vernalization requirements. However, old landraces from the Iberian Peninsula, which also have a vernalization requirement, possess this allele at a much higher frequency than modern winter barley cultivars. Under field conditions in which the vernalization requirement of winter cultivars is not satisfied, the dominant PPDH2 allele promotes flowering, even under increasing photoperiods above 12 h. This hypothesis was supported by expression analysis of vernalization-responsive genotypes. When the dominant allele of PPDH2 was expressed, this was associated with enhanced levels of VRNH1 and VRNH3 expression. Expression of these two genes is needed for the induction of flowering. Therefore, both in the field and under controlled conditions, PPDH2 has an effect of promotion of flowering.ConclusionsThe dominant, ancestral, allele of PPDH2 is prevalent in southern European barley germplasm. The presence of the dominant allele is associated with early expression of VRNH1 and early flowering. We propose that PPDH2 promotes flowering of winter cultivars under all non-inductive conditions, i.e. under short days or long days in plants that have not satisfied their vernalization requirement. This mechanism is indicated to be a component of an adaptation syndrome of barley to Mediterranean conditions.