We used a novel approach that incorporated chromosome sorting, next-generation sequencing, array hybridization, and systematic exploitation of conserved synteny with model grasses to assign ;86% of the estimated ;32,000 barley (Hordeum vulgare) genes to individual chromosome arms. Using a series of bioinformatically constructed genome zippers that integrate gene indices of rice (Oryza sativa), sorghum (Sorghum bicolor), and Brachypodium distachyon in a conserved synteny model, we were able to assemble 21,766 barley genes in a putative linear order. We show that the barley (H) genome displays a mosaic of structural similarity to hexaploid bread wheat (Triticum aestivum) A, B, and D subgenomes and that orthologous genes in different grasses exhibit signatures of positive selection in different lineages. We present an ordered, information-rich scaffold of the barley genome that provides a valuable and robust framework for the development of novel strategies in cereal breeding.

Phytophthora species secrete a large array of effectors during infection of their host plants. The Crinkler (CRN) gene family encodes a ubiquitous but understudied class of effectors with possible but as of yet unknown roles in infection. To appreciate CRN effector function in Phytophthora, we devised a simple Crn gene identification and annotation pipeline to improve effector prediction rates. We predicted 84 full-length CRN coding genes and assessed CRN effector domain diversity in sequenced Oomycete genomes. These analyses revealed evidence of CRN domain innovation in Phytophthora and expansion in the Peronosporales. We performed gene expression analyses to validate and define two classes of CRN effectors, each possibly contributing to infection at different stages. CRN localisation studies revealed that P. capsici CRN effector domains target the nucleus and accumulate in specific sub-nuclear compartments. Phenotypic analyses showed that few CRN domains induce necrosis when expressed in planta and that one cell death inducing effector, enhances P. capsici virulence on Nicotiana benthamiana. These results suggest that the CRN protein family form an important class of intracellular effectors that target the host nucleus during infection. These results combined with domain expansion in hemi-biotrophic and necrotrophic pathogens, suggests specific contributions to pathogen lifestyles. This work will bolster CRN identification efforts in other sequenced oomycete species and set the stage for future functional studies towards understanding CRN effector functions.

More than 2,000 genome-wide barley single nucleotide polymorphisms (SNPs) were developed by resequencing unigene fragments from eight diverse accessions. The average genome-wide SNP frequency observed in 877 unigenes was 1 SNP per 200 bp. However, SNP frequency was highly variable with the least number of SNP and SNP haplotypes observed within European cultivated germplasm reflecting effects of breeding history on genetic diversity. More than 300 SNP loci were mapped genetically in three experimental mapping populations which allowed the construction of an integrated SNP map incorporating a large number of RFLP, AFLP and SSR markers (1,237 loci in total). The genes used for SNP discovery were selected based on their transcriptional response to a variety of abiotic stresses. A set of known barley abiotic stress QTL was positioned on the linkage map, while the available sequence and gene expression information facilitated the identification of genes potentially associated with these traits. Comparison of the sequenced SNP loci to the rice genome sequence identified several regions of highly conserved gene order providing a framework for marker saturation in barley genomic regions of interest. The integration of genome-wide SNP and expression data with available genetic and phenotypic information will facilitate the identification of gene function in barley and other non-model organisms.

Although significant work has been undertaken regarding the response of model and crop plants to heat shock during the acclimatory phase, few studies have examined the steadystate response to the mild heat stress encountered in temperate agriculture. In the present work, we therefore exposed tuberizing potato plants to mildly elevated temperatures (30/ 20°C, day/night) for up to 5 weeks and compared tuber yield, physiological and biochemical responses, and leaf and tuber metabolomes and transcriptomes with plants grown under optimal conditions (22/16°C). Growth at elevated temperature reduced tuber yield despite an increase in net foliar photosynthesis. This was associated with major shifts in leaf and tuber metabolite profiles, a significant decrease in leaf glutathione redox state and decreased starch synthesis in tubers. Furthermore, growth at elevated temperature had a profound impact on leaf and tuber transcript expression with large numbers of transcripts displaying a rhythmic oscillation at the higher growth temperature. RT-PCR revealed perturbation in the expression of circadian clock transcripts including StSP6A, previously identified as a tuberization signal. Our data indicate that potato plants grown at moderately elevated temperatures do not exhibit classic symptoms of abiotic stress but that tuber development responds via a diversity of biochemical and molecular signals.

Bud break in raspberry (Rubus idaeus L.) is often poor and uneven, with many of the subapical buds remaining in a dormant state. In order to determine the dormancy status of raspberry buds, an empirical measure of bud burst in a growth-permissive environment following exposure to chilling (4 degrees C cold storage) was developed. For cv. Glen Ample, percentage bud burst in intact canes and isolated nodes was recorded after 14 d. Isolated nodes (a measure of endodormancy) achieved 100% bud burst after approximately 1500 h chilling whereas buds on intact plants (combined endo- and paradormancy) required an additional 1000 h chilling. A microarray approach was used to follow changes in gene expression that occurred during dormancy transition. The probes for the microarrays were obtained from endodormant and paradormant raspberry bud cDNA libraries. The expression profiles of 5300 clones from these libraries were subjected to principal component analysis to determine the most significant expression patterns. Sequence analysis of these clones, in many cases, enabled their functional categorization and the development of hypotheses concerning the mechanisms of bud dormancy release. Thus a set of novel candidates for key dormancy-related genes from raspberry buds have been identified. Bud dormancy is fundamental to the study of plant developmental processes and, in addition, its regulation is of significant economic importance to fruit and horticultural industries.

BackgroundPlant-microbe interactions feature complex signal interplay between pathogens and their hosts. Phytophthora species comprise a destructive group of fungus-like plant pathogens, collectively affecting a wide range of plants important to agriculture and natural ecosystems. Despite the availability of genome sequences of both hosts and microbes, little is known about the signal interplay between them during infection. In particular, accurate descriptions of coordinate relationships between host and microbe transcriptional programs are lacking.ResultsHere, we explore the molecular interaction between the hemi-biotrophic broad host range pathogen Phytophthora capsici and tomato. Infection assays and use of a composite microarray allowed us to unveil distinct changes in both P. capsici and tomato transcriptomes, associated with biotrophy and the subsequent switch to necrotrophy. These included two distinct transcriptional changes associated with early infection and the biotrophy to necrotrophy transition that may contribute to infection and completion of the P. capsici lifecycleConclusionsOur results suggest dynamic but highly regulated transcriptional programming in both host and pathogen that underpin P. capsici disease and hemi-biotrophy. Dynamic expression changes of both effector-coding genes and host factors involved in immunity, suggests modulation of host immune signaling by both host and pathogen. With new unprecedented detail on transcriptional reprogramming, we can now explore the coordinate relationships that drive host-microbe interactions and the basic processes that underpin pathogen lifestyles. Deliberate alteration of lifestyle-associated transcriptional changes may allow prevention or perhaps disruption of hemi-biotrophic disease cycles and limit damage caused by epidemics.

Dakota 58105-5677 (J.C.S.)The factors that regulate storage organ carotenoid content remain to be fully elucidated, despite the nutritional and economic importance of this class of compound. Recent findings suggest that carotenoid pool size is determined, at least in part, by the activity of carotenoid cleavage dioxygenases. The aim of this study was to investigate whether Carotenoid Cleavage Dioxygenase4 (CCD4) activity affects potato (Solanum tuberosum) tuber carotenoid content. Microarray analysis revealed elevated expression of the potato CCD4 gene in mature tubers from white-fleshed cultivars compared with higher carotenoid yellow-fleshed tubers. The expression level of the potato CCD4 gene was down-regulated using an RNA interference (RNAi) approach in stable transgenic lines. Down-regulation in tubers resulted in an increased carotenoid content, 2-to 5-fold higher than in control plants. The increase in carotenoid content was mainly due to elevated violaxanthin content, implying that this carotenoid may act as the in vivo substrate. Although transcript level was also reduced in plant organs other than tubers, such as leaves, stems, and roots , there was no change in carotenoid content in these organs. However, carotenoid levels were elevated in flower petals from RNAi lines. As well as changes in tuber carotenoid content, tubers from RNAi lines exhibited phenotypes such as heat sprouting, formation of chain tubers, and an elongated shape. These results suggest that the product of the CCD4 reaction may be an important factor in tuber heat responses.

The rhizosphere, the thin layer of soil surrounding and influenced by plant roots, defines a distinct and selective microbial habitat compared to unplanted soil. The microbial communities inhabiting the rhizosphere, the rhizosphere microbiota, engage in interactions with their host plants which span from parasitism to mutualism. Therefore, the rhizosphere microbiota emerges as one of the determinants of yield potential in crops. Studies conducted with different plant species have unequivocally pointed to the host plant as a driver of the microbiota thriving at the root–soil interface. Thus far, the host genetic traits shaping the rhizosphere microbiota are not completely understood. As root hairs play a critical role in resource exchanges between plants and the rhizosphere, we hypothesized that they can act as a determinant of the microbiota thriving at the root–soil interface. To test this hypothesis, we took advantage of barley (Hordeum vulgare) mutant lines contrasting for their root hair characteristics. Plants were grown in two agricultural soils, differentiating in their organic matter contents, under controlled environmental conditions. At early stem elongation rhizosphere specimens were collected and subjected to high-resolution 16S rRNA gene profiling. Our data revealed that the barley rhizosphere microbiota is largely dominated by members of the phyla Bacteroidetes and Proteobacteria, regardless of the soil type and the root hair characteristics of the host plant. Conversely, ecological indices calculated using operational taxonomic units (OTUs) presence, abundance, and phylogeny revealed a significant impact of root hair mutations on the composition of the rhizosphere microbiota. In particular, our data indicate that mutant plants host a reduced-complexity community compared to wild-type genotypes and unplanted soil controls. Congruently, the host genotype explained up to 18% of the variation in ecological distances computed for the rhizosphere samples. Importantly, this effect is manifested in a soil-dependent manner. A closer inspection of the sequencing profiles revealed that the root hair-dependent diversification of the microbiota is supported by a taxonomically narrow group of bacteria, with a bias for members of the orders Actinomycetales, Burkholderiales, Rhizobiales, Sphingomonadales, and Xanthomonadales. Taken together, our results indicate that the presence and function of root hairs are a determinant of the bacterial community thriving in the rhizosphere and their perturbations can markedly impact on the recruitment of individual members of the microbiota.