Strategies for Pyramiding Resistance Genes Against the Barley Yellow Mosaic Virus Complex (BaMMV,BaYMV, BaYMV-2)

Barley Yellow Mosaic Virus disease caused by different strains of BaYMV and BaMMV is a major threat to winter barley cultivation in Europe. Pyramiding of resistance genes may be considered as a promising strategy to avoid the selection of new virus strains and to create more durable resistances. However, this goal cannot be achieved by phenotypic selection due to the lack of differentiating virus strains. For pyramiding of resistance genes rym4, rym5, rym9 and rym11, located on chromosomes 3H and 4H of barley two different strategies have been developed. These strategies are based on doubled haploid lines (DHs) and marker assisted selection procedures. On the one hand F₁ derived DH-plants of single crosses were screened by molecular markers for genotypes being homozygous recessive for both resistance genes. These genotypes were crossed to lines carrying one resistance gene in common and an additional third gene, leading to a DH-population of which 25% carry three resistance genes, 50% have two resistance genes and 25% possess a single resistance gene homozygous recessively. Alternatively, F₁ plants having one resistance gene in common were directly inter-crossed [e.g. (rym4 × rym9) × (rym4 × rym11)] and about 100 seeds were produced per combination. Within these complex cross progenies plants were identified by markers being homozygous at the common resistance locus and heterozygous at the others. From such plants, theoretically present at a frequency of 6.25%, DH-lines were produced, which were screened for the presence of genotypes carrying three or two recessive resistance genes in a homozygous state. Besides DH-plants carrying all possible two-gene combinations, 20 DH-plants out of 107 analysed carrying rym4, rym9, and rym11 and 27 out of 187 tested carrying rym5, rym9, and rym11 homozygously have been detected using the second strategy which is faster but needs co-dominant markers, because in contrast to the first strategy marker selection is carried out on heterozygous genotypes.     

Direct targeting and rapid isolation of BAC clones spanning a defined chromosome region

To isolate genes of interest in plants, it is essential to construct bacterial artificial chromosome (BAC) libraries from specific genotypes. Construction and organisation of BAC libraries is laborious and costly, especially from organisms with large and complex genomes. In the present study, we developed the pooled BAC library strategy that allows rapid and low cost generation and screening of genomic libraries from any genotype of interest. The BAC library is constructed, directly organised into a few pools and screened for BAC clones of interest using PCR and hybridisation steps, without requiring organization into individual clones. As a proof of concept, a pooled BAC library of approximately 177,000 recombinant clones has been constructed from the barley cultivar Cebada Capa that carries the Rph7 leaf rust resistance gene. The library has an average insert size of 140 kb, a coverage of six barley genome equivalents and is organised in 138 pools of about 1,300 clones each. We rapidly established a single contig of six BAC clones spanning 230 kb at the Rph7 locus on chromosome 3HS. The described low-cost cloning strategy is fast and will greatly facilitate direct targeting of genes and large-scale intra- and inter-species comparative genome analysis.Edwige Isidore and Beatrice Scherrer contributed equally to the work.     

An alternative to radiation hybrid mapping for large-scale genome analysis in barley

The presence of a monosomic gametocidal chromosome (GC) in a barley chromosome addition line of common wheat generates structural aberrations in the barley chromosome as well as in the wheat chromosomes of gametes lacking the GC. A collection of structurally aberrant barley chromosomes is analogous to a panel of radiation hybrid (RH) mapping and is valuable for high-throughput physical mapping. We developed 90 common wheat lines (GC lines) containing aberrant barley 7H chromosomes induced by a gametocidal chromosome, 2C. DNAs isolated from these GC lines provided a panel of 7H chromosomal fragments in a wheat genetic background, comparable with RH mapping panels in mammals. We used this 7H GC panel and the methodology for RH mapping to physically map PCR-based barley markers, SSRs and AFLPs, onto chromosome 7H, relying on polymorphism between the 7H chromosome and the wheat genome. We call this method GC mapping. This study describes a novel adaptation and combination of methods of inducing chromosomal rearrangements to produce physical maps of markers. The advantages of the presented method are similar to RH mapping in that non-polymorphic markers can be used and the mapping panels can be relatively easily obtained. In addition, mapping results are cumulative when using the same mapping set with new markers. The GC lines will be available from the National Bioresources Project-KOMUGI (). Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

应用大麦杆控制原水蓝藻生长的初步研究

蓝藻对饮水安全有潜在威胁,但在水处理过程中很难除去。华南某地原水水塘多年来受到以假鱼腥藻类为主的蓝藻水华影响,尽管曾使用多种方法进行清除,但都只能起到应急的作用,因此需要寻求长期有效控制蓝藻生长的新方法。已有研究表明大麦杆在控制淡水藻类生长方面具有一定功效,且对环境无明显负面影响。为了解大麦杆对华南某地蓝藻,特别是假鱼腥藻类生长的影响,探讨在该地区气候条件下发挥抑藻作用所需时间和效果,为原水蓝藻控制措施的采取提供科学依据,自从2005年3月起至2005年7月,选用2个沉淀池作对比实验,实验池投入g·m-3的大麦杆,对照池不加大麦杆。研究表明大麦杆对原水中假鱼腥藻类的生长有较好的抑制作用,在该地气候条件下,最少需要一至两个月的时间,大麦杆才开始产生控藻作用。     

Barley anther culture: effects of annual cycle and spike position on microspore embryogenesis and albinism

The effect of donor plants annual cycle and anther/spike position on the production of microspore-derived plants and albinism were studied. We used the winter cv. Igri and the spring cv. Cork, known to respond similarly in anther culture but to produce 78% and 2% of green plants, respectively. In both cvs. the number of microspore-derived plants was significantly higher when the anthers were collected from January to July than from August to December. However, during this period the proportion of albino plants was not altered. Conversely, the anther response decreased from 76.6 to 31.5% in Igri and from 58.8 to 32.0% in Cork when the donor spike originates from the main shoot or the fourth tiller. Significantly, anthers collected from spike of the second tiller enabled us to drastically increase the proportion of regenerated green plantlets, by 16% in Igri and 1800% in Cork.     

Complex microcolinearity among wheat, rice, and barley revealed by fine mapping of the genomic region harboring a major QTL for resistance to Fusarium head blight in wheat

A major quantitative trait locus (QTL), Qfhs.ndsu-3BS, for resistance to Fusarium head blight (FHB) in wheat has been identified and verified by several research groups. The objectives of this study were to construct a fine genetic map of this QTL region and to examine microcolinearity in the QTL region among wheat, rice, and barley. Two simple sequence repeat (SSR) markers (Xgwm533 and Xgwm493) flanking this QTL were used to screen for recombinants in a population of 3,156 plants derived from a single F₇ plant heterozygous for the Qfhs.ndsu-3BS region. A total of 382 recombinants were identified, and they were genotyped with two more SSR markers and eight sequence-tagged site (STS) markers. A fine genetic map of the Qfhs.ndsu-3BS region was constructed and spanned 6.3 cM. Based on replicated evaluations of homozygous recombinant lines for Type II FHB resistance, Qfhs.ndsu-3BS, redesignated as Fhb1, was placed into a 1.2-cM marker interval flanked by STS3B-189 and STS3B-206. Primers of STS markers were designed from wheat expressed sequence tags homologous to each of six barley genes expected to be located near this QTL region. A comparison of the wheat fine genetic map and physical maps of rice and barley revealed inversions and insertions/deletions. This suggests a complex microcolinearity among wheat, rice, and barley in this QTL region.     

Cadmium-Induced Inhibition of Apoplastic Ascorbate Oxidase in Barley Roots

The effect of excess cadmium (Cd: 0.0, 0.25, 0.5, 1.0 and 2.0 mM) on growth and ascorbate oxidase (AO) activity was investigated in barley (Hordeum vulgare L. cv. Jubilant) roots. The study employed a filter-paper technique to germinate and grow the germinating seeds following imbibition with respective Cd treatments for 4 h at 25 °C in darkness. Cd was required at 1.0 mM to affect 50% root growth inhibition 72 h after the treatment. This Cd-induced root growth inhibition was accompanied by a corresponding loss of plasma membrane integrity in root cells as evaluated by Evans blue uptake. Excess Cd (1.0 and 2.0 mM) significantly inhibited the AO activity in all the analysed fractions of barley roots such as extracellular, soluble, cell wall (CW)- and membrane-bound fractions. AO was localized in the apoplast, and its highest specific activity was detected in the CW II fraction obtained by extraction with 1.0 M NaCl from purified cell walls. The analysis of AO isozyme profile showed that besides the reduced activity of two anionic and two cationic isozymes, one cationic AO isozyme was activated during excess Cd treatment, which could be detected in cell wall fractions CW II, III and IV.     

Haploid formation in maize, barley, flax, and potato

Summary. The article is reviewing some significant features and issues in the process of haploid formation in two important monocotyledonous crop plants – maize and barley – and in two dicotyledonous plants – flax and potato. Exotic maize lines with higher androgenic response turned up as a good source for this heritable trait and this valuable trait can be incorporated into elite maize lines via crossing. Lots of attempts were devoted to identifying some cytological and/or morphological markers for androgenic response in maize microspore cultures. The “starlike” organization of the cytoplasm inside the induced maize microspores together with the enlarged size of induced microspores can be considered as morphological markers for androgenic response. In barley, microspores with rich cytoplasm that was of granular appearance with the nucleus located near the cell wall and with no visible vacuole had the largest survival rate and many of these cells continued in development and produced embryos. In flax, a dramatic increase of induction rate in anther cultures (up to 25%) was achieved when flax anthers were pretreated for 3 days at 4 °C and afterwards kept for 1 day at 35 °C. Also gynogenesis in flax has been reported already and complete plants were obtained. In potato microspore cultures, formation of two dissimilar cells indicated a strong polarization in the system and as a result of this polarization a prominent suspensor developed that persisted until the torpedo stage of the androgenic embryo. This was the first time the formation of a well developed suspensor was described in connection with androgenesis. Correspondence and reprints: Institute of Plant Genetics and Biotechnology, Slovak Academy of Sciences, P.O. Box 39A, 950 07 Nitra, Slovak Republic.     

Plastid genome characterisation in Brassica and Brassicaceae using a new set of nine SSRs

The objective of the present study was to identify favourable exotic Quantitative Trait Locus (QTL) alleles for the improvement of agronomic traits in the BC₂DH population S42 derived from a cross between the spring barley cultivar Scarlett and the wild barley accession ISR42-8 (Hordeum vulgare ssp. spontaneum). QTLs were detected as a marker main effect and/or a marker × environment interaction effect (M × E) in a three-factorial ANOVA. Using field data of up to eight environments and genotype data of 98 SSR loci, we detected 86 QTLs for nine agronomic traits. At 60 QTLs the marker main effect, at five QTLs the M × E interaction effect, and at 21 QTLs both the effects were significant. The majority of the M × E interaction effects were due to changes in magnitude and are, therefore, still valuable for marker assisted selection across environments. The exotic alleles improved performance in 31 (36.0%) of 86 QTLs detected for agronomic traits. The exotic alleles had favourable effects on all analysed quantitative traits. These favourable exotic alleles were detected, in particular on the short arm of chromosome 2H and the long arm of chromosome 4H. The exotic allele on 4HL, for example, improved yield by 7.1%. Furthermore, the presence of the exotic allele on 2HS increased the yield component traits ears per m² and thousand grain weight by 16.4% and 3.2%, respectively. The present study, hence, demonstrated that wild barley does harbour valuable alleles, which can enrich the genetic basis of cultivated barley and improve quantitative agronomic traits.