BackgroundHigh density genetic maps of plants have, nearly without exception, made use of marker datasets containing missing or questionable genotype calls derived from a variety of genic and non-genic or anonymous markers, and been presented as a single linear order of genetic loci for each linkage group. The consequences of missing or erroneous data include falsely separated markers, expansion of cM distances and incorrect marker order. These imperfections are amplified in consensus maps and problematic when fine resolution is critical including comparative genome analyses and map-based cloning. Here we provide a new paradigm, a high-density consensus genetic map of barley based only on complete and error-free datasets and genic markers, represented accurately by graphs and approximately by a best-fit linear order, and supported by a readily available SNP genotyping resource.ResultsApproximately 22,000 SNPs were identified from barley ESTs and sequenced amplicons; 4,596 of them were tested for performance in three pilot phase Illumina GoldenGate assays. Data from three barley doubled haploid mapping populations supported the production of an initial consensus map. Over 200 germplasm selections, principally European and US breeding material, were used to estimate minor allele frequency (MAF) for each SNP. We selected 3,072 of these tested SNPs based on technical performance, map location, MAF and biological interest to fill two 1536-SNP "production" assays (BOPA1 and BOPA2), which were made available to the barley genetics community. Data were added using BOPA1 from a fourth mapping population to yield a consensus map containing 2,943 SNP loci in 975 marker bins covering a genetic distance of 1099 cM.ConclusionThe unprecedented density of genic markers and marker bins enabled a high resolution comparison of the genomes of barley and rice. Low recombination in pericentric regions is evident from bins containing many more than the average number of markers, meaning that a large number of genes are recombinationally locked into the genetic centromeric regions of several barley chromosomes. Examination of US breeding germplasm illustrated the usefulness of BOPA1 and BOPA2 in that they provide excellent marker density and sensitivity for detection of minor alleles in this genetically narrow material.

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Because of the huge size of the common wheat (Triticum aestivum L., 2n ϭ 6x ϭ 42, AABBDD) genome of 17,300 Mb, sequencing and mapping of the expressed portion is a logical first step for gene discovery. Here we report mapping of 7104 expressed sequence tag (EST) unigenes by Southern hybridization into a chromosome bin map using a set of wheat aneuploids and deletion stocks. Each EST detected a mean of 4.8 restriction fragments and 2.8 loci. More loci were mapped in the B genome (5774) than in the A (5173) or D (5146) genomes. The EST density was significantly higher for the D genome than for the A or B. In general, EST density increased relative to the physical distance from the centromere. The majority of EST-dense regions are in the distal parts of chromosomes. Most of the agronomically important genes are located in EST-dense regions. The chromosome bin map of ESTs is a unique resource for SNP analysis, comparative mapping, structural and functional analysis, and polyploid evolution, as well as providing a framework for constructing a sequence-ready, BAC-contig map of the wheat genome.

Dehydrins (DHNs; late embryogenesis abundant D-11) are a family of plant proteins induced in response to abiotic stresses such as drought, low temperature, and salinity or during the late stages of embryogenesis. Spectral and thermal properties of these proteins in purified form suggest that they are "intrinsically unstructured." However, DHNs contain at least one copy of a consensus 15-amino acid sequence, the "K segment," which resembles a class A2 amphipathic ␣-helical, lipid-binding domain found in other proteins such as apolipoproteins and ␣-synuclein. The presence of the K segment raises the question of whether DHNs bind lipids, bilayers, or phospholipid vesicles. Here, we show that maize (Zea mays) DHN DHN1 can bind to lipid vesicles that contain acidic phospholipids. We also observe that DHN1 binds more favorably to vesicles of smaller diameter than to larger vesicles, and that the association of DHN1 with vesicles results in an apparent increase of ␣-helicity of the protein. Therefore, DHNs, and presumably somewhat similar plant stress proteins in the late embryogenesis abundant and cold-regulated classes may undergo function-related conformational changes at the water/ membrane interface, perhaps related to the stabilization of vesicles or other endomembrane structures under stress conditions.When plant cells are under environmental stresses such as drought or low temperature, diverse physiological and molecular responses can occur, including alteration of gene expression, changes in metabolism, osmotic adjustment, induction of degradation and repair systems, and elevated expression of late embryogenesis abundant (LEA), chaperone, and mRNA-binding proteins (Ingram and Bartels, 1996; Thomashow, 1999).DHNs (LEA D-11 family) are among the most prevalent plant proteins induced during the late stage of embryogenesis or under drought, low temperature, freezing, salinity, or abscisic acid application (Close, 1996). They are characterized by diverse combinations of typical domains (Fig. 1A). The K segment is a Lys-rich 15-amino acid consensus sequence (EKKGIMDKIKEKPLG) that is highly conserved in all plants (Close, 1997). The K segment resembles a lipid-binding class A2 amphipathic ␣-helical segment found in apolipoproteins and ␣-synucleins ( Fig. 1B; Close, 1997; Davidson et al., 1998). The S-segment is a phosphorylatable Ser-rich tract. The Y segment is an N-terminal conserved sequence. The -segment is rich in polar amino acids and either Gly or a combination of Pro and Ala. Each DHN can be subclassified on the basis of these domains (Close, 1997).Immunolocalization studies of maize scutellar parenchyma cells have shown that DHN in the cytosol is associated with membrane-rich areas surrounding lipid and protein bodies (Asghar et al., 1994; EgertonWarburton et al., 1997), and in wheat (Triticum aestivum), an acidic DHN accumulates near the plasma membrane under cold stress (Danyluk et al., 1998). Despite this and other evidence of membrane binding in vivo, DHNs purified from these and numerous other sources...

Dehydrins are characterized by the consensus KIKEKLPG amino acid sequence found near the carboxy terminus, and usually repeated from one to many times within the protein. A synthetic peptide containing this consensus sequence was used to produce specific antibodies that recognize dehydrins in a wide range of plants. This range covered two families of monocots, viz. Gramineae (Hordeum vulgare L., Triticum aestivum L., Zea mays L., Oryza sativa L.) and Liliaceae (Allium sativa L.), and five families of dicots, Malvaceae (Gossypium hirsutum L.), Solanaceae (Lycopersicon esculentum L.), Brassicaceae (Raphanus sativus L.), Fabaceae (Vigna unguiculata L.), and Cucurbitaceae (Cucumis sativus L.). Two families of gymnosperms, Pinaceae (Pinus edulis Engelm.) and Ginkgoaceae (Ginkgo biloba L.), were also included. For several plants in which dehydrin cDNA and genomic clones have previously been characterized, it now appears that the dehydrin family of proteins is larger, and the regulation of dehydrin expression much more complex, than earlier studies have shown.

Dehydrins (DHNs; late embryogenesis abundant D11 family) are a family of intrinsically unstructured plant proteins that accumulate in the late stages of seed development and in vegetative tissues subjected to water deficit, salinity, low temperature, or abscisic acid treatment. We demonstrated previously that maize (Zea mays) DHNs bind preferentially to anionic phospholipid vesicles; this binding is accompanied by an increase in α-helicity of the protein, and adoption of α-helicity can be induced by sodium dodecyl sulfate. All DHNs contain at least one “K-segment,” a lysine-rich 15-amino acid consensus sequence. The K-segment is predicted to form a class A2 amphipathic α-helix, a structural element known to interact with membranes and proteins. Here, three K-segment deletion proteins of maize DHN1 were produced. Lipid vesicle-binding assays revealed that the K-segment is required for binding to anionic phospholipid vesicles, and adoption of α-helicity of the K-segment accounts for most of the conformational change of DHNs upon binding to anionic phospholipid vesicles or sodium dodecyl sulfate. The adoption of structure may help stabilize cellular components, including membranes, under stress conditions.

Recent advances in high-throughput genotyping have made it easier to combine information from different mapping populations into consensus genetic maps, which provide increased marker density and genome coverage compared to individual maps. Previously, a single nucleotide polymorphism (SNP)-based genotyping platform was developed and used to genotype 373 individuals in four barley (Hordeum vulgare L.) mapping populations. This led to a 2943 SNP consensus genetic map with 975 unique positions. In this work, we add data from six additional populations and more individuals from one of the original populations to develop an improved consensus map from 1133 individuals. A stringent and systematic analysis of each of the 10 populations was performed to achieve uniformity. This involved reexamination of the four populations included in the previous map. As a consequence, we present a robust consensus genetic map that contains 2994 SNP loci mapped to 1163 unique positions. The map spans 1137.3 cM with an average density of one marker bin per 0.99 cM. A novel application of the genotyping platform for gene detection allowed the assignment of 2930 genes to fl ow-sorted chromosomes or arms, confi rmed the position of 2545 SNP-mapped loci, added chromosome or arm allocations to an additional 370 SNP loci, and delineated pericentromeric regions for chromosomes 2H to 7H. Marker order has been improved and map resolution has been increased by almost 20%. These increased precision outcomes enable more optimized SNP selection for markerassisted breeding and support association genetic analysis and map-based cloning. It will also improve the anchoring of DNA sequence scaffolds and the barley physical map to the genetic map.

Dehydrins (DHNs; LEA D11) are one of the typical families of plant proteins that accumulate in response to dehydration, low temperature, osmotic stress or treatment with abscisic acid (ABA), or during seed maturation. We previously found that three genes encoding low-molecular-weight DHNs (Dhn1, Dhn2 and Dhn9) map within a 15-cM region of barley chromosome 5H that overlaps a QTL for winterhardiness, while other Dhn genes encoding low- and high-molecular-weight DHNs are located on chromosomes 3H, 4H and 6H. Here we examine the expression of specific Dhn genes under conditions associated with expression of the winterhardiness phenotype. Plants grown at 4 degrees C or in the field in Riverside, California developed similar, modest levels of freezing tolerance, coinciding with little low-MW Dhn gene activity. Dicktoo (the more tolerant cultivar) and Morex (the less tolerant) grown in Saskatoon, Canada expressed higher levels of expression of genes for low-MW DHNs than did the same cultivars in Riverside, with expression being higher in Dicktoo than Morex. Dehydration or freeze-thaw also evoked expression of genes for low MW DHNs, suggesting that the dehydration component of freeze-thaw in the field induces low expression of genes encoding low-MW DHNs. These observations are consistent with the hypothesis that the major chilling-induced DHNs help to prime plant cells for acclimation to more intense cold, which then involves adaptation to dehydration during freeze-thaw cycling. A role for chromosome 5H-encoded DHNs in acclimation to more intense cold seems possible, even though it is not the basis of the major heritable variation in winterhardiness within the Dicktoo x Morex population.