There is an increasing awareness that as a result of structural variation, a reference sequence representing a genome of a single individual is unable to capture all of the gene repertoire found in the species. A large number of genes affected by presence/absence and copy number variation suggest that it may contribute to phenotypic and agronomic trait diversity. Here we show by analysis of the Brassica oleracea pangenome that nearly 20% of genes are affected by presence/absence variation. Several genes displaying presence/absence variation are annotated with functions related to major agronomic traits, including disease resistance, flowering time, glucosinolate metabolism and vitamin biosynthesis.

We describe the construction of a reference genetic linkage map for the Brassica A genome, which will form the backbone for anchoring sequence contigs for the Multinational Brassica rapa Genome Sequencing Project. Seventy-eight doubled haploid lines derived from anther culture of the F(1) of a cross between two diverse Chinese cabbage (B. rapa ssp. pekinensis) inbred lines, 'Chiifu-401-42' (C) and 'Kenshin-402-43' (K) were used to construct the map. The map comprises a total of 556 markers, including 278 AFLP, 235 SSR, 25 RAPD and 18 ESTP, STS and CAPS markers. Ten linkage groups were identified and designated as R1-R10 through alignment and orientation using SSR markers in common with existing B. napus reference linkage maps. The total length of the linkage map was 1,182 cM with an average interval of 2.83 cM between adjacent loci. The length of linkage groups ranged from 81 to 161 cM for R04 and R06, respectively. The use of 235 SSR markers allowed us to align the A-genome chromosomes of B. napus with those of B. rapa ssp. pekinensis. The development of this map is vital to the integration of genome sequence and genetic information and will enable the international research community to share resources and data for the improvement of B. rapa and other cultivated Brassica species.

The Arabidopsis CAB2 promoter was used to define the terminal genomic targets that are subject to regulation by the circadian clock. An in vivo cab::luciferase bioluminescent marker was used to enable the assaying of the expression of chimeric constructs with unprecedented sensitivity and time resolution in living seedlings. Dissection of -322 to +1 of the CAB2 promoter has revealed several interesting features: it was demonstrated that the 323 bp fragment contains at least one strong general positive element. The positive element contains an ACGT core sequence specifically bound by a protein activity, termed CUF-1, and contributes to high level expression but is not required for phytochrome- or circadian-regulation. Moreover, a 78 bp domain was defined that confers both circadian- and phytochrome-regulation upon heterologous promoters. Conserved GATA sequences within the 78 bp regulatory domain are specifically bound by a protein factor designated CGF-1. The binding specificity of CGF-1 appears to be related to the GT-family of trihelix DNA-binding proteins. The role of these DNA-protein interactions is discussed in terms of clock- and phytochrome regulation, and their relevance as targets for pathways defined by photomorphogenic mutants.

Calcium (Ca) and magnesium (Mg) are the most abundant group II elements in both plants and animals. Genetic variation in shoot Ca and shoot Mg concentration (shoot Ca and Mg) in plants can be exploited to biofortify food crops and thereby increase dietary Ca and Mg intake for humans and livestock. We present a comprehensive analysis of within-species genetic variation for shoot Ca and Mg, demonstrating that shoot mineral concentration differs significantly between subtaxa (varietas). We established a structured diversity foundation set of 376 accessions to capture a high proportion of species-wide allelic diversity within domesticated Brassica oleracea, including representation of wild relatives (C genome, 1n 5 9) from natural populations. These accessions and 74 modern F 1 hybrid cultivars were grown in glasshouse and field environments. Shoot Ca and Mg varied 2-and 2.3-fold, respectively, and was typically not inversely correlated with shoot biomass, within most subtaxa. The closely related capitata (cabbage) and sabauda (Savoy cabbage) subtaxa consistently had the highest mean shoot Ca and Mg. Shoot Ca and Mg in glasshouse-grown plants was highly correlated with data from the field. To understand and dissect the genetic basis of variation in shoot Ca and Mg, we studied homozygous lines from a segregating B. oleracea mapping population. Shoot Ca and Mg was highly heritable (up to 40%). Quantitative trait loci (QTL) for shoot Ca and Mg were detected on chromosomes C2, C6, C7, C8, and, in particular, C9, where QTL accounted for 14% to 55% of the total genetic variance. The presence of QTL on C9 was substantiated by scoring recurrent backcross substitution lines, derived from the same parents. This also greatly increased the map resolution, with strong evidence that a 4-cM region on C9 influences shoot Ca. This region corresponds to a 0.41-Mb region on Arabidopsis (Arabidopsis thaliana) chromosome 5 that includes 106 genes. There is also evidence that pleiotropic loci on C8 and C9 affect shoot Ca and Mg. Map-based cloning of these loci will reveal how shoot-level phenotypes relate to Ca 21 and Mg 21 uptake and homeostasis at the molecular level.

Brassica napus , an allotetraploid crop, is hypothesized to be a hybrid from unknown varieties of Brassica rapa and Brassica oleracea . Despite the economic importance of B. napus , much is unresolved regarding its phylogenomic relationships, genetic structure, and diversification. Here we conduct a comprehensive study among diverse accessions from 183 B. napus (including rapeseed, rutabaga, and Siberian kale), 112 B. rapa , and 62 B. oleracea and its wild relatives. Using RNA-seq of B. napus accessions, we define the genetic diversity and sub-genome variance of six genetic clusters. Nuclear and organellar phylogenies for B. napus and its progenitors reveal varying patterns of inheritance and post-formation introgression. We discern regions with signatures of selective sweeps and detect 8,187 differentially expressed genes with implications for B. napus diversification. This study highlights the complex origin and evolution of B. napus providing insights that can further facilitate B. napus breeding and germplasm preservation.