Trait analysis reveals DOG1 determines initial depth of seed dormancy, but not changes during dormancy cycling that result in seedling emergence timing.

Seedling emergence timing is crucial in competitive plant communities and so contributes to species fitness. To understand the mechanistic basis of variation in seedling emergence timing, we exploited the contrasting behaviour of two Arabidopsis thaliana ecotypes: Cape Verde Islands (Cvi) and Burren (Bur-0). We used RNA-Seq analysis of RNA from exhumed seeds and quantitative trait loci (QTL) analyses on a mapping population from crossing the Cvi and Bur-0 ecotypes. We determined genome-wide expression patterns over an annual dormancy cycle in both ecotypes, identifying nine major clusters based on the seasonal timing of gene expression, and variation in behaviour between them. QTL were identified for depth of seed dormancy and seedling emergence timing (SET). Both analyses showed a key role for DOG1 in determining depth of dormancy, but did not support a direct role for DOG1 in generating altered seasonal patterns of seedling emergence. The principle QTL determining SET (SET1: dormancy cycling) is physically close on chromosome 5, but is distinct from DOG1. We show that SET1 and two other SET QTLs each contain a candidate gene (AHG1, ANAC060, PDF1 respectively) closely associated with DOG1 and abscisic acid signalling and suggest a model for the control of SET in the field.

Assembly and characterisation of a unique onion diversity set identifies resistance to Fusarium basal rot and improved seedling vigour.

KEY MESSAGE: A unique, global onion diversity set was assembled, genotyped and phenotyped for beneficial traits. Accessions with strong basal rot resistance and increased seedling vigour were identified along with associated markers. Conserving biodiversity is critical for safeguarding future crop production. Onion (Allium cepa L.) is a globally important crop with a very large (16 Gb per 1C) genome which has not been sequenced. While onions are self-fertile, they suffer from severe inbreeding depression and as such are highly heterozygous as a result of out-crossing. Bulb formation is driven by daylength, and accessions are adapted to the local photoperiod. Onion seed is often directly sown in the field, and hence seedling establishment is a critical trait for production. Furthermore, onion yield losses regularly occur worldwide due to Fusarium basal rot caused by Fusarium oxysporum f. sp. cepae. A globally relevant onion diversity set, consisting of 10 half-sib families for each of 95 accessions, was assembled and genotyping carried out using 892 SNP markers. A moderate level of heterozygosity (30-35%) was observed, reflecting the outbreeding nature of the crop. Using inferred phylogenies, population structure and principal component analyses, most accessions grouped according to local daylength. A high level of intra-accession diversity was observed, but this was less than inter-accession diversity. Accessions with strong basal rot resistance and increased seedling vigour were identified along with associated markers, confirming the utility of the diversity set for discovering beneficial traits. The onion diversity set and associated trait data therefore provide a valuable resource for future germplasm selection and onion breeding.

Genome-Wide Linkage and Association Mapping of Halo Blight Resistance in Common Bean to Race 6 of the Globally Important Bacterial Pathogen.

Pseudomonas syringae pv. phaseolicola (Psph) Race 6 is a globally prevalent and broadly virulent bacterial pathogen with devastating impact causing halo blight of common bean (Phaseolus vulgaris L.). Common bean lines PI 150414 and CAL 143 are known sources of resistance against this pathogen. We constructed high-resolution linkage maps for three recombinant inbred populations to map resistance to Psph Race 6 derived from the two common bean lines. This was complemented with a genome-wide association study (GWAS) of Race 6 resistance in an Andean Diversity Panel of common bean. Race 6 resistance from PI 150414 maps to a single major-effect quantitative trait locus (QTL; HB4.2) on chromosome Pv04 and confers broad-spectrum resistance to eight other races of the pathogen. Resistance segregating in a Rojo × CAL 143 population maps to five chromosome arms and includes HB4.2. GWAS detected one QTL (HB5.1) on chromosome Pv05 for resistance to Race 6 with significant influence on seed yield. The same HB5.1 QTL, found in both Canadian Wonder × PI 150414 and Rojo × CAL 143 populations, was effective against Race 6 but lacks broad resistance. This study provides evidence for marker-assisted breeding for more durable halo blight control in common bean by combining alleles of race-nonspecific resistance (HB4.2 from PI 150414) and race-specific resistance (HB5.1 from cv. Rojo).

Population Structure of Sclerotinia subarctica and Sclerotinia sclerotiorum in England, Scotland and Norway.

Sclerotinia species are important fungal pathogens of a wide range of crops and wild host plants. While the biology and population structure of Sclerotinia sclerotiorum has been well-studied, little information is available for the related species S. subarctica. In this study, Sclerotinia isolates were collected from different crop plants and the wild host Ranuculus ficaria (meadow buttercup) in England, Scotland, and Norway to determine the incidence of Sclerotinia subarctica and examine the population structure of this pathogen for the first time. Incidence was very low in England, comprising only 4.3% of isolates while moderate and high incidence of S. subarctica was identified in Scotland and Norway, comprising 18.3 and 48.0% of isolates respectively. Characterization with eight microsatellite markers identified 75 haplotypes within a total of 157 isolates over the three countries with a few haplotypes in Scotland and Norway sampled at a higher frequency than the rest across multiple locations and host plants. In total, eight microsatellite haplotypes were shared between Scotland and Norway while none were shared with England. Bayesian and principal component analyses revealed common ancestry and clustering of Scottish and Norwegian S. subarctica isolates while English isolates were assigned to a separate population cluster and exhibited low diversity indicative of isolation. Population structure was also examined for S. sclerotiorum isolates from England, Scotland, Norway, and Australia using microsatellite data, including some from a previous study in England. In total, 484 haplotypes were identified within 800 S. sclerotiorum isolates with just 15 shared between England and Scotland and none shared between any other countries. Bayesian and principal component analyses revealed a common ancestry and clustering of the English and Scottish isolates while Norwegian and Australian isolates were assigned to separate clusters. Furthermore, sequencing part of the intergenic spacer (IGS) region of the rRNA gene resulted in 26 IGS haplotypes within 870 S. sclerotiorum isolates, nine of which had not been previously identified and two of which were also widely distributed across different countries. S. subarctica therefore has a multiclonal population structure similar to S. sclerotiorum, but has a different ancestry and distribution across England, Scotland, and Norway.

Towards new sources of resistance to the currant-lettuce aphid (Nasonovia ribisnigri).

Domesticated lettuce varieties encompass much morphological variation across a range of crop type groups, with large collections of cultivars and landrace accessions maintained in genebanks. Additional variation not captured during domestication, present in ancestral wild relatives, represents a potentially rich source of alleles that can deliver to sustainable crop production. However, these large collections are difficult and costly to screen for many agronomically important traits. In this paper, we describe the generation of a diversity collection of 96 lettuce and wild species accessions that are amenable to routine phenotypic analysis and their genotypic characterization with a panel of 682 newly developed expressed sequence tag (EST)-linked KASP™ single nucleotide polymorphism (SNP) markers that are anchored to the draft Lactuca sativa genome assembly. To exemplify the utility of these resources, we screened the collection for putative sources of resistance to currant-lettuce aphid (Nasonovia ribisnigri) and carried out association analyses to look for potential SNPs linked to resistance.

Trait to gene analysis reveals that allelic variation in three genes determines seed vigour.

Predictable seedling establishment is essential for resource-efficient and cost-effective crop production; it is widely accepted as a critically important trait determining yield and profitability. Seed vigour is essential to this, but its genetic basis is not understood. We used natural variation and fine mapping in the crop Brassica oleracea to show that allelic variation at three loci influence the key vigour trait of rapid germination. Functional analysis in both B. oleracea and the model Arabidopsis identified and demonstrated activity of genes at these loci. Two candidate genes were identified at the principal Speed of Germination QTL (SOG1) in B. oleracea. One gene BoLCVIG2 is a homologue of the alternative-splicing regulator (AtPTB1). The other gene BoLCVIG1 was unknown, but different alleles had different splice forms that were coincident with altered abscisic acid (ABA) sensitivity. We identified a further QTL, Reduced ABscisic Acid 1 (RABA1) that influenced ABA content and provide evidence that this results from the activity of a homologue of the ABA catabolic gene AtCYP707A2 at this locus. Lines containing beneficial alleles of these three genes had greater seed vigour. We propose a mechanism in which both seed ABA content and sensitivity to it determines speed of germination.

Transcriptome and methylome profiling reveals relics of genome dominance in the mesopolyploid Brassica oleracea.

BACKGROUND: Brassica oleracea is a valuable vegetable species that has contributed to human health and nutrition for hundreds of years and comprises multiple distinct cultivar groups with diverse morphological and phytochemical attributes. In addition to this phenotypic wealth, B. oleracea offers unique insights into polyploid evolution, as it results from multiple ancestral polyploidy events and a final Brassiceae-specific triplication event. Further, B. oleracea represents one of the diploid genomes that formed the economically important allopolyploid oilseed, Brassica napus. A deeper understanding of B. oleracea genome architecture provides a foundation for crop improvement strategies throughout the Brassica genus. RESULTS: We generate an assembly representing 75% of the predicted B. oleracea genome using a hybrid Illumina/Roche 454 approach. Two dense genetic maps are generated to anchor almost 92% of the assembled scaffolds to nine pseudo-chromosomes. Over 50,000 genes are annotated and 40% of the genome predicted to be repetitive, thus contributing to the increased genome size of B. oleracea compared to its close relative B. rapa. A snapshot of both the leaf transcriptome and methylome allows comparisons to be made across the triplicated sub-genomes, which resulted from the most recent Brassiceae-specific polyploidy event. CONCLUSIONS: Differential expression of the triplicated syntelogs and cytosine methylation levels across the sub-genomes suggest residual marks of the genome dominance that led to the current genome architecture. Although cytosine methylation does not correlate with individual gene dominance, the independent methylation patterns of triplicated copies suggest epigenetic mechanisms play a role in the functional diversification of duplicate genes.

Genetic regulation of glucoraphanin accumulation in Beneforté broccoli.

· Diets rich in broccoli (Brassica oleracea var italica) have been associated with maintenance of cardiovascular health and reduction in risk of cancer. These health benefits have been attributed to glucoraphanin that specifically accumulates in broccoli. The development of broccoli with enhanced concentrations of glucoraphanin may deliver greater health benefits. · Three high-glucoraphanin F1 broccoli hybrids were developed in independent programmes through genome introgression from the wild species Brassica villosa. Glucoraphanin and other metabolites were quantified in experimental field trials. Global SNP analyses quantified the differential extent of B. villosa introgression · The high-glucoraphanin broccoli hybrids contained 2.5-3 times the glucoraphanin content of standard hybrids due to enhanced sulphate assimilation and modifications in sulphur partitioning between sulphur-containing metabolites. All of the high-glucoraphanin hybrids possessed an introgressed B. villosa segment which contained a B. villosa Myb28 allele. Myb28 expression was increased in all of the high-glucoraphanin hybrids. Two high-glucoraphanin hybrids have been commercialised as Beneforté broccoli. · The study illustrates the translation of research on glucosinolate genetics from Arabidopsis to broccoli, the use of wild Brassica species to develop cultivars with potential consumer benefits, and the development of cultivars with contrasting concentrations of glucoraphanin for use in blinded human intervention studies.

High-resolution mapping of a fruit firmness-related quantitative trait locus in tomato reveals epistatic interactions associated with a complex combinatorial locus.

Fruit firmness in tomato (Solanum lycopersicum) is determined by a number of factors including cell wall structure, turgor, and cuticle properties. Firmness is a complex polygenic trait involving the coregulation of many genes and has proved especially challenging to unravel. In this study, a quantitative trait locus (QTL) for fruit firmness was mapped to tomato chromosome 2 using the Zamir Solanum pennellii interspecific introgression lines (ILs) and fine-mapped in a population consisting of 7,500 F2 and F3 lines from IL 2-3 and IL 2-4. This firmness QTL contained five distinct subpeaks, Fir(s.p.)QTL2.1 to Fir(s.p.)QTL2.5, and an effect on a distal region of IL 2-4 that was nonoverlapping with IL 2-3. All these effects were located within an 8.6-Mb region. Using genetic markers, each subpeak within this combinatorial locus was mapped to a physical location within the genome, and an ethylene response factor (ERF) underlying Fir(s.p.)QTL2.2 and a region containing three pectin methylesterase (PME) genes underlying Fir(s.p.)QTL2.5 were nominated as QTL candidate genes. Statistical models used to explain the observed variability between lines indicated that these candidates and the nonoverlapping portion of IL 2-4 were sufficient to account for the majority of the fruit firmness effects. Quantitative reverse transcription-polymerase chain reaction was used to quantify the expression of each candidate gene. ERF showed increased expression associated with soft fruit texture in the mapping population. In contrast, PME expression was tightly linked with firm fruit texture. Analysis of a range of recombinant lines revealed evidence for an epistatic interaction that was associated with this combinatorial locus.