Comparative AB-QTL analysis in barley using a single exotic donor of Hordeum vulgare ssp. spontaneum.

This paper reports on the first comparative advanced backcross quantitative trait locus (AB-QTL) study in barley. The BC₂F₂ population H×101 was generated from crossing var. Harry [H; Hordeum vulgare ssp. vulgare (Hv)] with ISR101-23 [101; H. v. ssp. spontaneum (Hsp)]. The results of the AB-QTL analysis for 13 quantitative traits are presented and, subsequently, compared with the AB-QTL study of the barley cross Apex × ISR101-23 (A×101; Pillen et al., Theor Appl Genet 107:340–352). Both AB populations share the exotic Hsp donor accession ISR101-23. In H×101, 108 putative QTLs (17%) were identified among the 650 marker×trait combinations tested. Altogether 52 (48 %) favorable effects were identified from the exotic parent. At these marker loci, the homozygous Hsp genotype was associated with an improvement in the trait compared to the homozygous Hv genotype. The percentage of QTLs detected in H×101 was comparable to that in A×101 (17% vs. 15%), however more favorable exotic QTL alleles were located in H×101 than in A×101 (48% vs. 34%). In H×101, the Hsp QTL allele at EBmac0679_[4H] was associated with a yield increase of 5.9% averaged across the six environments tested. A comparison of putative QTLs between H×101 and A×101 was based on 26 shared SSR markers. In total, 26% of the putative QTLs could be detected simultaneously in both AB populations. This finding indicates that only a portion of the QTL effects of the donor allele can be transferred from one elite recipient to the next.Communicated by G. WenzelThis work is dedicated to the 65th birthday of Prof. Dr. H.H. Geiger, Department of Plant Breeding, Seed Science and Population Genetics, University of Hohenheim, Germany.     

Characterization of genetic diversity in core collection accessions of wild barley,Hordeum vulgare ssp. spontaneum

Genetic variability in the 143 core accessions of wild barley, Hordeum vulgare ssp. spontaneum, was assessed by allozyme analysis. A total of 34 alleles were detected at ten isozyme loci. All loci were polymorphic except Pgd-1, which was monomorphic. Est-2 and Est-4 were the most diverse loci, with genetic diversity values of 0.747 and 0.686, respectively. The comparison of the results with those of previous studies indicates that all alleles occurring in cultivated and wild barley are observed in this set of the wild Barley Core Collection. Only one allele (Pgd-1 Tj) was absent. It is noteworthy that one new allele at the Ndh-2 locus and another new allele at Aco-2 locus were first detected in the present study. Nine of the 34 alleles were rare and detected only in one to four accessions. The genetic similarities among the 143 accessions ranged from 0.18 to 1.00. Data analysis based on clustering and principal coordinate analysis showed that a high level of genetic variability exists in this set of core accessions, and indicated that some duplication probably exists in this set core based on the present study.     

Measuring root traits in barley (Hordeum vulgare ssp. vulgare and ssp. spontaneum) seedlings using gel chambers,soil sacs and X-ray microtomography

Plant and Soil - Many sites inside a protected area in Apulia region (Italy) have been contaminated with heavy metals (Cd, Cr, Cu, Ni, Pb, Zn) because an inadequate disposal of a variety of wastes...     

Analysis of chromosomal polymorphism in barley (Hordeum vulgare L. ssp. vulgare) and between H. vulgare and H. chilense using three-color fluorescence in situ hybridization (FISH)

The aim of the present work was to study chromosomal polymorphism within cultivated barley (Hordeum vulgare ssp. vulgare) using three-color fluorescence in situ hybridization (FISH). The physical distribution of the most frequently used, highly repetitive DNA sequences (GAA)₇ specific for pericentromeric heterochromatic regions, the ribosomal DNA clone pTa71, specific for the 45S rDNA, and the barley-specific telomere-associated sequence HvT01, was investigated to reveal genetic diversity in metaphase spreads of ten barley genotypes with diverse geographical origin, growth habit and row number. A wild relative of barley, Hordeum chilense was also studied in order to compare the polymorphism between and within Hordeum species. Significant differences in the hybridization patterns of all three DNA probes could be detected between the two related species, but only probes pTa71 and HvT01 showed variation in the intensity and/or position of hybridization sites among genotypes of H. vulgare ssp. vulgare. The extent of polymorphism was less than that earlier reported for molecular markers and was restricted to the long chromosome arms, with differences between the chromosomes. 1H and 3H proved to be the most variable chromosomes and 4H and 6H the most conserved.     

Localisation of genes for resistance against Blumeria graminis f.sp. hordei and Puccinia graminis in a cross between a barley cultivar and a wild barley (Hordeum vulgare ssp. spontaneum) line

The aims of this investigation have been to map new (quantitative) resistance genes against powdery mildew, caused by Blumeria graminis f.sp. hordei L., and leaf rust, caused by Puccinia hordei L., in a cross between the barley ( Hordeum vulgare ssp. vulgare) cultivar "Vada" and the wild barley ( Hordeum vulgare ssp. spontaneum) line "1B-87" originating from Israel. The population consisted of 121 recombinant inbred lines. Resistance against leaf rust and powdery mildew was tested on detached leaves. The leaf rust isolate "I-80" and the powdery mildew isolate "Va-4", respectively, were used for the infection in this experiment. Moreover, powdery mildew disease severity was observed in the field at two different epidemic stages. In addition to other DNA markers, the map included 13 RGA (resistance gene analog) loci. The structure of the data demanded a non-parametric QTL-analysis. For each of the four observations, two QTLs with very high significance were localised. QTLs for resistance against powdery mildew were detected on chromosome 1H, 2H, 3H, 4H and 7H. QTLs for resistance against leaf rust were localised on 2H and 6H. Only one QTL was common for two of the powdery mildew related traits. Three of the seven QTLs were localised at the positions of the RGA-loci. Three of the five powdery mildew related QTLs are sharing their chromosomal position with known qualitative resistance genes. All detected QTLs behaved additively. Possible sources of the distorted segregation observed, the differences between the results for the different powdery mildew related traits and the relation between qualitative and quantitative resistance are discussed.     

The influence of linkage and inbreeding on patterns of nucleotide sequence diversity at duplicate alcohol dehydrogenase loci in wild barley (Hordeum vulgare ssp. spontaneum)

Patterns of nucleotide sequence diversity are analyzed for three duplicate alcohol dehydrogenase loci (adh1-adh3) within a species-wide sample of 25 accessions of wild barley (Hordeum vulgare ssp. spontaneum). The adh1 and adh2 loci are tightly linked (recombination fraction <0.01) while the adh3 locus is inherited independently. Wild barley is predominantly self-fertilizing (approximately 98%), and as a consequence, effective recombination is restricted by the extreme reduction in heterozygosity. Large reductions in effective recombination, in turn, widen the conditions for linkage to influence nucleotide sequence diversity through the action of selective sweeps or background selection. These considerations would appear to predict (1) homogeneity in patterns of nucleotide sequence diversity, especially between closely linked loci, and (2) extensive linkage disequilibrium relative to random-mating species. In contrast to these expectations, the wild barley data reveal heterogeneity in patterns of nucleotide sequence diversity and levels of linkage disequilibrium that are indistinguishable from those observed at adh1 in maize, an outbreeding grass species.     

Physical geography,isolation by distance and environmental variables shape genomic variation of wild barley (Hordeum vulgare L. ssp. spontaneum) in the Southern Levant

Determining the extent of genetic variation that reflects local adaptation in crop-wild relatives is of interest for the purpose of identifying useful genetic diversity for plant breeding. We investigated the association of genomic variation with geographical and environmental factors in wild barley (Hordeum vulgare L. ssp. spontaneum) populations of the Southern Levant using genotyping by sequencing (GBS) of 244 accessions in the Barley 1K+ collection. The inference of population structure resulted in four genetic clusters that corresponded to eco-geographical habitats and a significant association between lower gene flow rates and geographical barriers, e.g. the Judaean Mountains and the Sea of Galilee. Redundancy analysis (RDA) revealed that spatial autocorrelation explained 45% and environmental variables explained 15% of total genomic variation. Only 4.5% of genomic variation was solely attributed to environmental variation if the component confounded with spatial autocorrelation was excluded. A synthetic environmental variable combining latitude, solar radiation, and accumulated precipitation explained the highest proportion of genomic variation (3.9%). When conditioned on population structure, soil water capacity was the most important environmental variable explaining 1.18% of genomic variation. Genome scans with outlier analysis and genome-environment association studies were conducted to identify adaptation signatures. RDA and outlier methods jointly detected selection signatures in the pericentromeric regions, which have reduced recombination, of the chromosomes 3H, 4H, and 5H. However, selection signatures mostly disappeared after correction for population structure. In conclusion, adaptation to the highly diverse environments of the Southern Levant over short geographical ranges had a limited effect on the genomic diversity of wild barley. This highlighted the importance of nonselective forces in genetic differentiation.Subject terms: Genetic variation, Agriculture, Evolutionary ecology, Evolutionary genetics, Plant evolution     

Heritable somaclonal variation in wild barley (Hordeum spontaneum)

Summary Progenies of H. spontaneum plants regenerated from immature embryo derived calli were analysed for somaclonal variation on the following traits: (1) organization of the intergenic spacer of the rRNA genes; (2) B and C hordein pattern on SDS-PAGE; (3) genomic organization of the B and C hordein coding sequences; (4) mitochondrial DNA organization assayed by hybridization of Southern blots of total DNA with mitochondrial coding genes; (5) cytology. One out of twelve progeny plants was characterized as variant for two traits: (a) a loss of 1.8 and 2.5 kb Taq I intergenic rDNA spacer fragments and (b) a variant pattern of hordeins on 1-D SDS-PAGE. No numerical or structural chromosome variation was detected among the control plants therefore it is assumed that the variation was caused by the in vitro culture and transmitted, through sexual reproduction, to the analysed progeny.     

Genetic analysis of the accumulation of COR14 proteins in wild (Hordeum spontaneum) and cultivated (Hordeum vulgare) barley

The cold-regulated (COR14) protein of 14 kDa is a polypeptide accumulated under low-temperature conditions in the chloroplasts of barley leaves. In H. vulgare the COR14 antibody cross-reacts with two proteins, with a slightly different relative molecular weight around the marker of 14.4 kDa, referred to as COR14a and COR14b (high and low relative molecular weight, respectively). In a collection of H. spontaneum genotypes a clear polymorphism was found for the corresponding COR proteins. While some accessions showed the same COR pattern as cultivated barley, in 38 out of 61 accessions examined the COR14 antibody cross-reacted with an additional coldregulated protein with a relative molecular weight of about 24 kDa (COR24). The accumulation of COR24 was often associated with the absence of COR14b; the relationship between the COR14b/COR24 polymorphism and the adaptation of H. spontaneum to different environments is discussed. By studying COR14 accumulation in cultivated barley we have found that the threshold induction-temperature of COR14a is associated with the loci controlling winter hardiness. This association was demonstrated by using either a set of 30 cultivars of different origin, or two sets of frost-tolerant and frost-sensitive F1 doubled-haploid lines derived from the cross Dicktoo (winter type) x Morex (spring type). These results suggest that the threshold induction-temperature of COR14a can be a potential biochemical marker for the identification of superior frostresistant barley genotypes.     

The essential oil of Origanum vulgare L. ssp. vulgare growing wild in vilnius district (Lithuania)

The plants of wild Origanum vulgare L. ssp. vulgare were collected in 10 localities of Vilnius district (Lithuania) in 1995-1999. The main constituents of the essential oils from 8 localities were beta-ocimene (14.9-21.6%), germacrene D (10.0-16.2), beta-caryophyllene (10.8- 15.7%) and sabinene (6.6- 4.2%). The essential oils from two localities contained only three above compounds as major components: germacrene D, beta-ocimene and sabinene or beta-caryophyllene, beta-ocimene and germacrene D. Three chemotypes of essential oils were identified. The main chemotype was beta-ocimene germacrene D-beta-caryophyllene. The terpenic hydrocarbons made up 52.8-80.6% of the essential oils. The 42 identified components made up 85.6-98.0% of the essential oil.     

AFLP genetic polymorphism in wild barley (Hordeum spontaneum) populations in Israel

The genetic diversity produced by the amplified fragment length polymorphism (AFLP) method was studied in 94 genotypes of wild barley, Hordeum spontaneum (C. Koch) Thell., originating from ten ecologically and geographically different locations in Israel. Eight primer pairs produced 204 discernible loci of which 189 (93%) were polymorphic. Each genotype had a unique banding profile and the genetic similarity coefficient varied between 0.74 and 0.98. The phenogram generated from these similarities by the UPGMA method did not group genotypes strictly according to their geographical origin, which pattern was also seen in the principal coordinate (PCO) plot. Genetic diversity was larger within (69%) than among (31%) populations. Associations between ecogeographical variables and the mean gene diversity were found at one primer pair. The results are discussed and compared with data obtained by the simple sequence repeat (SSR) method.     

AB-QTL analysis in spring barley. I. Detection of resistance genes against powdery mildew, leaf rust and scald introgressed from wild barley

The objective of this study was to map new resistance genes against powdery mildew (Blumeria graminis f. sp. hordei L.), leaf rust (Puccinia hordei L.) and scald [Rhynchosporium secalis (Oud.) J. Davis] in the advanced backcross doubled haploid (BC₂DH) population S42 derived from a cross between the spring barley cultivar Scarlett and the wild barley accession ISR42-8 (Hordeum vulgare ssp. spontaneum). Using field data of disease severity recorded in eight environments under natural infestation and genotype data of 98 SSR loci, we detected nine QTL for powdery mildew, six QTL for leaf rust resistance and three QTL for scald resistance. The presence of the exotic QTL alleles reduced disease symptoms by a maximum of 51.5, 37.6 and 16.5% for powdery mildew, leaf rust and scald, respectively. Some of the detected QTL may correspond to previously identified qualitative (i.e. Mla) and to quantitative resistance genes. Others may be newly identified resistance genes. For the majority of resistance QTL (61.0%) the wild barley contributed the favourable allele demonstrating the usefulness of wild barley in the quest for resistant cultivars.     

Detection and quantification of Paenibacillus polymyxa in the rhizosphere of wild barley (Hordeum spontaneum) with real-time PCR

Aim:  To detect and quantify the plant drought tolerance enhancing bacterium Paenibacillus polymyxa in a collection of 160 Hordeum spontaneum rhizosphere samples at the 'Evolution Canyon' ('EC'), Israel.
Methods and Results:  PCR primers and a FAM-TAMRA probe (6-carboxyfluorescein, 6-carboxy-tetramethyl-rhodamine) targeting 16S rRNA genes were designed and used to detect and quantify the target strain. Two commercial kits, Bio101 Fast Spin and Mo Bio Ultra Clean Soil DNA, were tested for DNA isolation from the rhizosphere and surrounding soil. Population densities of P. polymyxa were studied in the rhizosphere of wild barley and surrounding soil from the contrasting climatic slopes at the 'EC' using the real-time PCR and culture based methods.
Conclusion:  Paenibacillus polymyxa is one of the best established species in wild barley rhizosphere at the 'EC' slopes. With the real-time PCR assay we are able to detect 1 pg of DNA per PCR corresponding to 100 cells per ml. The results at the 'EC' correlate well to bacterial estimations by culture based methods.
Significance and Impact of the Study:  Significantly higher P. polymyxa cell number was detected in the rhizosphere of arid 'African' microclimate indicating possible role of adaptive co-evolution with plants.     

Haploid plants regenerated via anther culture in wild barley (Hordeum spontaneum C. Kock)

Summary Androgenic plants have been obtained via anther culture in four natural populations of Hordeum spontaneum. Microscopic observations revealed that androgenesis started with the formation of two vegetative-type nuclei derived from the mitotic division of the uninucleate microspores. In this species androgenesis was affected by the type and concentration of the sugars added to the culture medium: the highest response (17% of callusing anthers) was observed on media containing 80 g l^–1 maltose. The highest production of androgenic plants (per 100 anthers, 5.9 green and 4.3 albino plants) was obtained from callus grown on these same media. About half of the green plants regenerated were haploid, while the others were diploid and set seed.Abbreviations IAA indolacetic acid - BAP 6-benzylaminopurine     

Grain protein variability among populations of wild barley (Hordeum spontaneum C. Koch.) from Jordan

Summary Protein content, kernel weight, and genetic diversity in the storage protein hordein, encoded by the Hor 1 and Hor 2 loci, were assessed in 12 populations of wild barley (Hordeum spontaneum C. Koch.) collected from central, peripheral, and marginal areas of its distribution in Jordan. Protein content ranged from 106.3 to 239.1 g kg^-1, and kernel weight ranged from 21.17 to 31.8 mg. Populations with high protein content and heavy kernels have been identified. Electrophoretic analysis of the storage protein hordein showed that the two hordein loci, Hor 1 and Hor 2, are highly polymorphic, having 34 and 38 alleles, respectively. Polymorphism (H_e) was highest in central populations (H_e Hor 1=0.859, H_e Hor 2=0.782), intermediate in peripheral populations (H_e Hor 1=0.566, H_e Hor 2=0.509), and lowest in marginal populations (H_e Hor 1=0.392, H_e Hor 2=0.349). Geographical distances between populations were not indicative of Nei's genetic similarity (NI). NI values averaged 0.209 and ranged from 0.0 to 0.83, supporting the hypothesis of an island population model for the species. The high proportion of allelic diversity, apportioned among populations for Hor 1 (0.584) and Hor 2 (0.495) loci, indicates that these natural populations are a rich reserve of genetic variability for protein. This variability is readily exploitable in breeding.     

Microgeographic edaphic differentiation in allozyme polymorphisms of wild barley (Hordeum spontaneum,Poaceae)

Allozymic variation in proteins encoded by 22 loci was analyzed electrophoretically in 278 individual plants of wild barley,Hordeum spontaneum, the progenitor of cultivated barley, in four 100 meter transects, in Israel, each equally subdivided into basalt and terra rossa soil types. Significant differentiation according to soil was found in 9 alleles. Our results suggest that allozyme polymorphisms in wild barley are at least partly adaptive, and differentiate by edaphic natural selection rather than by stochastic processes, and/or neutrality of allozymic variants.     

Development and characterization of recombinant chromosome substitution lines (RCSLs) using Hordeum vulgare subsp. spontaneum as a source of donor alleles in a Hordeum vulgare subsp. vulgare background.

The ancestor of barley (Hordeum vulgare subsp. spontaneum) may be a source of novel alleles for crop improvement. We developed a set of recombinant chromosome substitution lines (RCSLs) using an accession of H. vulgare subsp. spontaneum (Caesarea 26-24, from Israel) as the donor and Hordeum vulgare subsp. vulgare 'Harrington' (the North American malting quality standard) as the recurrent parent via two backcrosses to the recurrent parent, followed by six generations of selfing. Here we report (i) the genomic architecture of the RCSLs, as inferred by simple sequence repeat (SSR) markers, and (ii) the effects of H. vulgare subsp. spontaneum genome segment introgressions in terms of three classes of phenotypes: inflorescence yield components, malting quality traits, and domestication traits. Significant differences among the RCSLs were detected for all phenotypes measured. The phenotypic effects of the introgressions were assessed using association analysis, and these were referenced to quantitative trait loci (QTL) reported in the literature. Hordeum vulgare subsp. spontaneum, despite its overall inferior phenotype, contributed some favorable alleles for agronomic and malting quality traits. In most cases, the introgression of the ancestral genome resulted in a loss of desirable phenotypes in the cultivated parent. Although disappointing from a plant breeding perspective, this finding may prove to be a useful tool for gene discovery.