Zoltán Bedo

Genetic analysis of the components of winterhardiness in barley (Hordeum vulgare L.)

Winterhardiness in cereals is the consequence of a number of complex and interacting component characters: cold tolerance, vernalization requirement, and photoperiod sensitivity. An understanding of the genetic basis of these component traits should allow for more-effective selection. Genome map-based analyses hold considerable promise for dissecting complex phenotypes. A 74-point linkage map was developed from 100 doubled haploid lines derived from a winter x spring barley cross and used as the basis for quantitative trait locus (QTL) analyses to determine the chromosome location of genes controlling components of winterhardiness. Despite the greater genome coverage provided by the current map, a previously-reported interval on chromosome 7 remains the only region where significant QTL effects for winter survival were detected in this population. QTLs for growth habit and heading date, under 16 h and 24 h light, map to the same region. A QTL for heading date under these photoperiod regimes also maps to chromosome 2. Contrasting alleles at these loci interact in an epistatic fashion. A distinct set of QTLs mapping to chromosomes 1, 2, 3, and 5 determined heading date under 8 h of light. Under field conditions, all QTLs identified under controlled environment conditions were determinants of heading date. Patterns of differential QTL expression, coupled with additive and additive x additive QTL effects, underscore the complexity of winterhardiness. The presence of unique phenotype combinations in the mapping population suggests that coincident QTLs for heading date and winter survival represent the effects of linkage rather than pleiotropy.

Phytochemicals and dietary fiber components in rye varieties in the HEALTHGRAIN diversity screen

Ten rye varieties grown in one location were analyzed for their contents of dietary fiber (arabinoxylan and beta-glucan) and phytochemicals (folate, tocols, phenolic acids, alkylresorcinols, and sterols). The varieties included old and modern varieties from five European countries. Significant differences were observed in the contents of all phytochemicals in whole grains and in the fiber contents in the flour and bran. The old French varieties Haute Loire and Queyras had high contents of most phytochemicals, whereas the Polish varieties Dankowskie-Zlote and Warko were relatively poor in phytochemicals. The varieties with a high content of folate tended to have low alkylresorcinol contents and vice versa. Furthermore, high contents of arabinoxylans were associated with high contents in tocols and sterols. The 10 selected rye samples comprising old populations and old and modern varieties from different ecological regions of Europe demonstrate high natural variation in their composition and show that landraces and old populations are useful genetic resources for plant breeding. The contents of single phytochemicals can likely be affected by breeding, and they may be adjusted by the right selection of genotype.

Variation in the Content of Dietary Fiber and Components Thereof in Wheats in the HEALTHGRAIN Diversity Screen

Within the HEALTHGRAIN diversity screening program, the variation in the content of dietary fiber and components thereof in different types of wheat was studied. The wheat types were winter (131 varieties) and spring (20 varieties) wheats (both Triticum aestivum L., also referred to as common wheats), durum wheat (Triticum durum Desf., 10 varieties), spelt wheat (Triticum spelta L., 5 varieties), einkorn wheat (T. monococcum L., 5 varieties), and emmer wheat (Triticum dicoccum Schubler, 5 varieties). Common wheats contained, on average, the highest level of dietary fiber [11.5-18.3% of dry matter (dm)], whereas einkorn and emmer wheats contained the lowest level (7.2-12.8% of dm). Intermediate levels were measured in durum and spelt wheats (10.7-15.5% of dm). Also, on the basis of the arabinoxylan levels in bran, the different wheat types could be divided this way, with ranges of 12.7-22.1% of dm for common wheats, 6.1-14.4% of dm for einkorn and emmer wheats, and 10.9-13.9% of dm for durum and spelt wheats. On average, bran arabinoxylan made up ca. 29% of the total dietary fiber content of wheat. In contrast to what was the case for bran, the arabinoxylan levels in flour were comparable between the different types of wheat. For wheat, in general, they varied between 1.35 and 2.75% of dm. Einkorn, emmer, and durum wheats contained about half the level of mixed-linkage beta-glucan (0.25-0.45% of dm) present in winter, spring, and spelt wheats (0.50-0.95% of dm). All wheat types had Klason lignin, the levels of which varied from 1.40 to 3.25% of dm. The arabinoxylan contents in bran and the dietary fiber contents in wholemeal were inversely and positively related with bran yield, respectively. Aqueous wholemeal extract viscosity, a measure for the level of soluble dietary fiber, was determined to large extent by the level of water-extractable arabinoxylan. In conclusion, the present study revealed substantial variation in the contents of dietary fiber and constituents thereof between different wheat types and varieties.

The HEALTHGRAIN Wheat Diversity Screen: Effects of Genotype and Environment on Phytochemicals and Dietary Fiber Components

Analysis of the contents of bioactive components (tocols, sterols, alkylresorcinols, folates, phenolic acids, and fiber components) in 26 wheat cultivars grown in six site x year combinations showed that the extent of variation due to variety and environment differed significantly between components. The total contents of tocols, sterols, and arabinoxylan fiber were highly heritable and hence an appropriate target for plant breeding. However, significant correlations between the contents of bioactive components and environmental factors (precipitation and temperature) during grain development also occurred, with even highly heritable components differing in amount between grain samples grown in different years on different sites.

Folate in Wheat Genotypes in the HEALTHGRAIN Diversity Screen

As part of the diversity screen of the HEALTHGRAIN project, the total folate contents of bread wheat (130 winter and 20 spring wheat genotypes), durum wheat (10 genotypes), earlier cultivated diploid einkorn and tetraploid emmer wheat (5 genotypes of each), and spelt (5 genotypes), grown in the same location in a controlled manner, were determined by a microbiological assay. The total folate contents ranged from 364 to 774 ng/g of dm in winter wheat and from 323 to 741 ng/g of dm in spring wheat, thus showing a marked variation. The highest mean for total folate content was measured in the durum wheat genotypes, whereas the earlier cultivated diploid and tetraploid wheat genotypes and spelt were shown to possess comparable or even higher folate contents than bread wheat. HPLC analysis of selected genotypes showed that 5-formyltetrahydrofolate was the major vitamer. The data provide a basis for breeding wheat genotypes with improved folate content.

Production of novel allelic variation for genes involved in starch biosynthesis through mutagenesis

Given the important role that starch plays in food and non-food uses of many crops, particularly wheat, efforts are being made to manipulate its composition through modification of the amylose/amylopectin ratio. Approaches used to achieve this goal include the manipulation of the genes involved in the starch biosynthetic pathway using natural or induced mutations and transgenic methods. The use of mutagenesis to produce novel allelic variation represents a powerful tool to increase genetic diversity and this approach seems particularly appropriate for starch synthase genes for which limited variation exists. In this work, an EMS-mutagenised population of bread wheat cv. Cadenza has been screened by combining SDS–PAGE analysis of granule bound starch proteins with a TILLING (Targeting Induced Local Lesions IN Genomes) approach at the gene level. In particular we have focused on two groups of synthase genes, those encoding the starch synthase II (Sgp-1) and those corresponding to the waxy proteins (Wx). SDS–PAGE analysis of granule bound proteins allowed the identification of single null genotypes associated with each of the three homoeologous loci. Molecular characterization of induced mutants has been performed using genome specific primer pairs for Sgp-1 and Wx genes. Additional novel allelic variation has also been detected at the different Sgp-1 homoeoloci by using a reverse genetic approach (TILLING). In particular single nucleotide substitutions, introducing a premature stop codon and creating amino acid substitutions, have been identified.

Natural variation in grain composition of wheat and related cereals

The wheat grain comprises three groups of major components, starch, protein, and cell wall polysaccharides (dietary fiber), and a range of minor components that may confer benefits to human health. Detailed analyses of dietary fiber and other bioactive components were carried out under the EU FP6 HEALTHGRAIN program on 150 bread wheat lines grown on a single site, 50 lines of other wheat species and other cereals grown on the same site, and 23-26 bread wheat lines grown in six environments. Principal component analysis allowed the 150 bread wheat lines to be classified on the basis of differences in their contents of bioactive components and wheat species (bread, durum, spelt, emmer, and einkorn wheats) to be clearly separated from related cereals (barley, rye, and oats). Such multivariate analyses could be used to define substantial equivalence when novel (including transgenic) cereals are considered.

Effects of Genotype and Environment on the Content and Composition of Phytochemicals and Dietary Fiber Components in Rye in the HEALTHGRAIN Diversity Screen

The effects of genotype and environment on the content of bioactive components in rye were determined with four varieties being grown on one site for three years and on three additional sites in the third year and a fourth variety being included in all trials except year 1. Clear differences were observed in the extent to which the contents of dietary fiber components (arabinoxylan, beta-glucan, total dietary fiber) and phytochemicals (folates, alkylresorcinols, sterols, tocols, phenolic acids) varied between varieties and between the same varieties grown in different sites (United Kingdom, France, Hungary, Poland) and years (2005-2007 in Hungary), with sterols being the most stable and phenolic acids the least. However, no single variety could be selected as having the highest overall level of bioactive components or as being more stable in comparison across environments.

Combined meta-genomics analyses unravel candidate genes for the grain dietary fiber content in bread wheat (Triticum aestivum L.)

Grain dietary fiber content in wheat not only affects its end use and technological properties including milling, baking and animal feed but is also of great importance for health benefits. In this study, integration of association genetics (seven detected loci on chromosomes 1B, 3A, 3D, 5B, 6B, 7A, 7B) and meta-QTL (three consensus QTL on chromosomes 1B, 3D and 6B) analyses allowed the identification of seven chromosomal regions underlying grain dietary fiber content in bread wheat. Based either on a diversity panel or on bi-parental populations, we clearly demonstrate that this trait is mainly driven by a major locus located on chromosome 1B associated with a log of p value >13 and a LOD score >8, respectively. In parallel, we identified 73 genes differentially expressed during the grain development and between genotypes with contrasting grain fiber contents. Integration of quantitative genetics and transcriptomic data allowed us to propose a short list of candidate genes that are conserved in the rice, sorghum and Brachypodium chromosome regions orthologous to the seven wheat grain fiber content QTL and that can be considered as major candidate genes for future improvement of the grain dietary fiber content in bread wheat breeding programs.

Bánkúti 1201--an old Hungarian wheat variety with special storage protein composition.

Abstract Bánkúti 1201, an old Hungarian wheat variety with special quality traits, was analysed to determine the relationships between its storage protein composition and superior quality-attributes for breadmaking. Based on the storage protein composition, the variety appears to have the nature of a population, containing several genotypes with different gluten protein alleles. Using molecular markers, a new mutant x-type HMW glutenin allele was identified, containing an extra cysteine residue and showing a moderate, positive-effect on gluten properties. In lines possessing subunits Bx7+By8 the overexpression of the Bx-type subunit could be detected, resulting in a higher unextractable polymeric protein (UPP) content and increased dough strength. It was found that the presence or absence of subunit Bx7 has an equilibrating effect on the dough extensibility, which is generally characteristic of the Bánkúti 1201 population. The complex good bread-making quality of the variety, which has strong but highly extensible dough, is probably due to the balance between lines which express subunit Bx7 and those which do not.