Single nucleotide polymorphism mapping and alignment of recombinant chromosome substitution lines in barley

Single nucleotide polymorphism (SNP) genotyping is useful for assessing genetic variation in germplasm collections, genetic map development and detection of alien chromosome substitutions. In this study, a diversity analysis using 1,301 SNPs on a set of 37 barley accessions was conducted. This analysis showed a high polymorphism rate between the malting barley cultivar 'Haruna Nijo' and the food barley cultivar 'Akashinriki'. Haruna Nijo and Akashinriki are donors of the barley expressed sequence tag (EST) collections. A doubled haploid (DH) population derived from the cross between Haruna Nijo and Akashinriki was genotyped with 1,448 SNPs. Of these 1,448 SNPs, 734 were polymorphic and distributed on barley linkage groups (chromosomes) as follows: 1H (86), 2H (125), 3H (120), 4H (100), 5H (127), 6H (88) and 7H (88). By using cMAP, we integrated the SNP markers across high-density maps. The SNPs were also used to genotype 98 BC(3)F(4) recombinant chromosome substitution lines (RCSLs) developed from the same cross (Haruna Nijo/Akashinriki). These data were used to create graphical genotypes for each line and thus estimate the location, extent and total number of introgressions from Akashinriki in the Haruna Nijo background. The 35 selected RCSLs sample most of the Akashinriki food barley genome, with only a few missing segments. These resources bring new alleles into the malting barley gene pool from food barley.     

Comprehensive sequence analysis of 24,783 barley full-length cDNAs derived from 12 clone libraries

Full-length cDNA (FLcDNA) libraries consisting of 172,000 clones were constructed from a two-row malting barley cultivar (Hordeum vulgare 'Haruna Nijo') under normal and stressed conditions. After sequencing the clones from both ends and clustering the sequences, a total of 24,783 complete sequences were produced. By removing duplicates between these and publicly available sequences, 22,651 representative sequences were obtained: 17,773 were novel barley FLcDNAs, and 1,699 were barley specific. Highly conserved genes were found in the barley FLcDNA sequences for 721 of 881 rice (Oryza sativa) trait genes with 50% or greater identity. These FLcDNA resources from our Haruna Nijo cDNA libraries and the full-length sequences of representative clones will improve our understanding of the biological functions of genes in barley, which is the cereal crop with the fourth highest production in the world, and will provide a powerful tool for annotating the barley genome sequences that will become available in the near future.     

Assembly and analysis of a qingke reference genome demonstrate its close genetic relation to modern cultivated barley

Qingke, the local name of hulless barley in the Tibetan Plateau, is a staple food for Tibetans. The availability of its reference genome sequences could be useful for studies on breeding and molecular evolution. Taking advantage of the third‐generation sequencer (PacBio), we de novo assembled a 4.84‐Gb genome sequence of qingke, cv. Zangqing320 and anchored a 4.59‐Gb sequence to seven chromosomes. Of the 46,787 annotated ‘high‐confidence’ genes, 31 564 were validated by RNA‐sequencing data of 39 wild and cultivated barley genotypes with wide genetic diversity, and the results were also confirmed by nonredundant protein database from NCBI. As some gaps in the reference genome of Morex were covered in the reference genome of Zangqing320 by PacBio reads, we believe that the Zangqing320 genome provides the useful supplements for the Morex genome. Using the qingke genome as a reference, we conducted a genome comparison, revealing a close genetic relationship between a hulled barley (cv. Morex) and a hulless barley (cv. Zangqing320), which is strongly supported by the low‐diversity regions in the two genomes. Considering the origin of Morex from its breeding pedigree, we then demonstrated a close genomic relationship between modern cultivated barley and qingke. Given this genomic relationship and the large genetic diversity between qingke and modern cultivated barley, we propose that qingke could provide elite genes for barley improvement.     

A whole-genome snapshot of 454 sequences exposes the composition of the barley genome and provides evidence for parallel evolution of genome size in wheat and barley

The genomes of barley and wheat, two of the world's most important crops, are very large and complex due to their high content of repetitive DNA. In order to obtain a whole-genome sequence sample, we performed two runs of 454 (GS20) sequencing on genomic DNA of barley cv. Morex, which yielded approximately 1% of a haploid genome equivalent. Almost 60% of the sequences comprised known transposable element (TE) families, and another 9% represented novel repetitive sequences. We also discovered high amounts of low-complexity DNA and non-genic low-copy DNA. We identified almost 2300 protein coding gene sequences and more than 660 putative conserved non-coding sequences. Comparison of the 454 reads with previously published genomic sequences suggested that TE families are distributed unequally along chromosomes. This was confirmed by in situ hybridizations of selected TEs. A comparison of these data for the barley genome with a large sample of publicly available wheat sequences showed that several TE families that are highly abundant in wheat are absent from the barley genome. This finding implies that the TE composition of their genomes differs dramatically, despite their very similar genome size and their close phylogenetic relationship.     

Analysis of Intraspecies Diversity in Wheat and Barley Genomes Identifies Breakpoints of Ancient Haplotypes and Provides Insight into the Structure of Diploid and Hexaploid Triticeae Gene Pools

A large number of wheat (Triticum aestivum) and barley (Hordeum vulgare) varieties have evolved in agricultural ecosystems since domestication. Because of the large, repetitive genomes of these Triticeae crops, sequence information is limited and molecular differences between modern varieties are poorly understood. To study intraspecies genomic diversity, we compared large genomic sequences at the Lr34 locus of the wheat varieties Chinese Spring, Renan, and Glenlea, and diploid wheat Aegilops tauschii. Additionally, we compared the barley loci Vrs1 and Rym4 of the varieties Morex, Cebada Capa, and Haruna Nijo. Molecular dating showed that the wheat D genome haplotypes diverged only a few thousand years ago, while some barley and Ae. tauschii haplotypes diverged more than 500,000 years ago. This suggests gene flow from wild barley relatives after domestication, whereas this was rare or absent in the D genome of hexaploid wheat. In some segments, the compared haplotypes were very similar to each other, but for two varieties each at the Rym4 and Lr34 loci, sequence conservation showed a breakpoint that separates a highly conserved from a less conserved segment. We interpret this as recombination breakpoints of two ancient haplotypes, indicating that the Triticeae genomes are a heterogeneous and variable mosaic of haplotype fragments. Analysis of insertions and deletions showed that large events caused by transposable element insertions, illegitimate recombination, or unequal crossing over were relatively rare. Most insertions and deletions were small and caused by template slippage in short homopolymers of only a few base pairs in size. Such frequent polymorphisms could be exploited for future molecular marker development.     

The genome sequence of silkworm, Bombyx mori.

We performed threefold shotgun sequencing of the silkworm (Bombyx mori) genome to obtain a draft sequence and establish a basic resource for comprehensive genome analysis. By using the newly developed RAMEN assembler, the sequence data derived from whole-genome shotgun (WGS) sequencing were assembled into 49,345 scaffolds that span a total length of 514 Mb including gaps and 387 Mb without gaps. Because the genome size of the silkworm is estimated to be 530 Mb, almost 97% of the genome has been organized in scaffolds, of which 75% has been sequenced. By carrying out a BLAST search for 50 characteristic Bombyx genes and 11,202 non-redundant expressed sequence tags (ESTs) in a Bombyx EST database against the WGS sequence data, we evaluated the validity of the sequence for elucidating the majority of silkworm genes. Analysis of the WGS data revealed that the silkworm genome contains many repetitive sequences with an average length of <500 bp. These repetitive sequences appear to have been derived from truncated transposons, which are interspersed at 2.5- to 3-kb intervals throughout the genome. This pattern suggests that silkworm may have an active mechanism that promotes removal of transposons from the genome. We also found evidence for insertions of mitochondrial DNA fragments at 9 sites. A search for Bombyx orthologs to Drosophila genes controlling sex determination in the WGS data revealed 11 Bombyx genes and suggested that the sex-determining systems differ profoundly between the two species.     

Detection of QTLs controlling alpha-amylase activity in a diversity panel of 343 barley accessions

The α–amylase activity of cultivated barley is critically important to the brewing industry. Here, we surveyed variation in malt α–amylase activity in 343 cultivated barley accessions from around the world. Population structure analysis based on genotype data at 1536 SNPs clustered these accessions into two groups, one comprising South-East Asian and Ethiopian accessions and one group containing the other accessions. A genome-wide association study identified significant quantitative trait loci (QTLs) for α–amylase activity on all seven chromosomes of barley. Accessions showing high and low α–amylase activity were crossed with the high-quality Japanese malting barley cv. Harun Nijo to develop F₂ mapping populations. We identified two QTLs on chromosome 6H in a cross between Haruna Nijo (high activity) × Weal (highest activity). Single QTLs were identified each on 3H, 4H, and 5H from a cross between Haruna Nijo (high activity) × VLB-1 (low activity), indicating that the high α–amylase activity in Haruna Nijo might be derived from loci on these chromosomes. The addition of the high α–amylase activity QTL alleles from chromosome 6H in cv. Weal further increased the α–amylase activity conferred by alleles of Haruna Nijo. These results demonstrate that a target haplotype can be successfully improved using a strategy comprising diversity analysis of ex situ collections followed by introducing effective new alleles.     

Sequencing of 15 622 gene‐bearing BACs clarifies the gene‐dense regions of the barley genome

Barley (Hordeum vulgare L.) possesses a large and highly repetitive genome of 5.1 Gb that has hindered the development of a complete sequence. In 2012, the International Barley Sequencing Consortium released a resource integrating whole‐genome shotgun sequences with a physical and genetic framework. However, because only 6278 bacterial artificial chromosome (BACs) in the physical map were sequenced, fine structure was limited. To gain access to the gene‐containing portion of the barley genome at high resolution, we identified and sequenced 15 622 BACs representing the minimal tiling path of 72 052 physical‐mapped gene‐bearing BACs. This generated ~1.7 Gb of genomic sequence containing an estimated 2/3 of all Morex barley genes. Exploration of these sequenced BACs revealed that although distal ends of chromosomes contain most of the gene‐enriched BACs and are characterized by high recombination rates, there are also gene‐dense regions with suppressed recombination. We made use of published map‐anchored sequence data from Aegilops tauschii to develop a synteny viewer between barley and the ancestor of the wheat D‐genome. Except for some notable inversions, there is a high level of collinearity between the two species. The software HarvEST:Barley provides facile access to BAC sequences and their annotations, along with the barley–Ae. tauschii synteny viewer. These BAC sequences constitute a resource to improve the efficiency of marker development, map‐based cloning, and comparative genomics in barley and related crops. Additional knowledge about regions of the barley genome that are gene‐dense but low recombination is particularly relevant.     

Sugarcane genome sequencing by methylation filtration provides tools for genomic research in the genus Saccharum

Many economically important crops have large and complex genomes that hamper their sequencing by standard methods such as whole genome shotgun (WGS). Large tracts of methylated repeats occur in plant genomes that are interspersed by hypomethylated gene‐rich regions. Gene‐enrichment strategies based on methylation profiles offer an alternative to sequencing repetitive genomes. Here, we have applied methyl filtration with McrBC endonuclease digestion to enrich for euchromatic regions in the sugarcane genome. To verify the efficiency of methylation filtration and the assembly quality of sequences submitted to gene‐enrichment strategy, we have compared assemblies using methyl‐filtered (MF) and unfiltered (UF) libraries. The use of methy filtration allowed a better assembly by filtering out 35% of the sugarcane genome and by producing 1.5× more scaffolds and 1.7× more assembled Mb in length compared with unfiltered dataset. The coverage of sorghum coding sequences (CDS) by MF scaffolds was at least 36% higher than by the use of UF scaffolds. Using MF technology, we increased by 134× the coverage of gene regions of the monoploid sugarcane genome. The MF reads assembled into scaffolds that covered all genes of the sugarcane bacterial artificial chromosomes (BACs), 97.2% of sugarcane expressed sequence tags (ESTs), 92.7% of sugarcane RNA‐seq reads and 98.4% of sorghum protein sequences. Analysis of MF scaffolds from encoded enzymes of the sucrose/starch pathway discovered 291 single‐nucleotide polymorphisms (SNPs) in the wild sugarcane species, S. spontaneum and S. officinarum. A large number of microRNA genes was also identified in the MF scaffolds. The information achieved by the MF dataset provides a valuable tool for genomic research in the genus Saccharum and for improvement of sugarcane as a biofuel crop.     

Construction of a virtual Mycobacterium tuberculosis consensus genome and its application to data from a next generation sequencer

Background

Although Mycobacterium tuberculosis isolates are consisted of several different lineages and the epidemiology analyses are usually assessed relative to a particular reference genome, M. tuberculosis H37Rv, which might introduce some biased results. Those analyses are essentially based genome sequence information of M. tuberculosis and could be performed in sillico in theory, with whole genome sequence (WGS) data available in the databases and obtained by next generation sequencers (NGSs). As an approach to establish higher resolution methods for such analyses, whole genome sequences of the M. tuberculosis complexes (MTBCs) strains available on databases were aligned to construct virtual reference genome sequences called the consensus sequence (CS), and evaluated its feasibility in in sillico epidemiological analyses.

Results

The consensus sequence (CS) was successfully constructed and utilized to perform phylogenetic analysis, evaluation of read mapping efficacy, which is crucial for detecting single nucleotide polymorphisms (SNPs), and various MTBC typing methods virtually including spoligotyping, VNTR, Long sequence polymorphism and Beijing typing. SNPs detected based on CS, in comparison with H37Rv, were utilized in concatemer-based phylogenetic analysis to determine their reliability relative to a phylogenetic tree based on whole genome alignment as the gold standard. Statistical comparison of phylogenic trees based on CS with that of H37Rv indicated the former showed always better results that that of later. SNP detection and concatenation with CS was advantageous because the frequency of crucial SNPs distinguishing among strain lineages was higher than those of H37Rv. The number of SNPs detected was lower with the consensus than with the H37Rv sequence, resulting in a significant reduction in computational time. Performance of each virtual typing was satisfactory and accorded with those published when those are available.

Conclusions

These results indicated that virtual CS constructed from genome sequence data is an ideal approach as a reference for MTBC studies.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1368-9) contains supplementary material, which is available to authorized users.     

Comparing whole‐genome shotgun sequencing and DNA metabarcoding approaches for species identification and quantification of pollen species mixtures

Molecular identification of mixed‐species pollen samples has a range of applications in various fields of research. To date, such molecular identification has primarily been carried out via amplicon sequencing, but whole‐genome shotgun (WGS) sequencing of pollen DNA has potential advantages, including (1) more genetic information per sample and (2) the potential for better quantitative matching. In this study, we tested the performance of WGS sequencing methodology and publicly available reference sequences in identifying species and quantifying their relative abundance in pollen mock communities. Using mock communities previously analyzed with DNA metabarcoding, we sequenced approximately 200Mbp for each sample using Illumina HiSeq and MiSeq. Taxonomic identifications were based on the Kraken k‐mer identification method with reference libraries constructed from full‐genome and short read archive data from the NCBI database. We found WGS to be a reliable method for taxonomic identification of pollen with near 100% identification of species in mixtures but generating higher rates of false positives (reads not identified to the correct taxon at the required taxonomic level) relative to rbcL and ITS2 amplicon sequencing. For quantification of relative species abundance, WGS data provided a stronger correlation between pollen grain proportion and sequence read proportion, but diverged more from a 1:1 relationship, likely due to the higher rate of false positives. Currently, a limitation of WGS‐based pollen identification is the lack of representation of plant diversity in publicly available genome databases. As databases improve and costs drop, we expect that eventually genomics methods will become the methods of choice for species identification and quantification of mixed‐species pollen samples.     

Chicken genome sequence: a centennial gift to poultry genetics

A draft sequence of the chicken genome will be available by early 2004. This event conveniently marks the start of the second century of poultry genetics, coming 100 years after the use of the chicken to demonstrate Mendelian inheritance in animals by William Bateson. How will the second, post-genomic century of poultry genetics differ from the first? A whole genome shotgun (WGS) approach is being used to obtain the chicken sequence, with the goal of generating approximately six-fold coverage of the genome. Bacterial artificial chromosome (BAC) and fosmid clone end sequences, along with a BAC contig map integrated with genetic linkage and radiation hybrid maps, will form the platform for assembly of the WGS data. Rapid progress in global analysis of chicken gene expression patterns is also being made. Comparative genomics will link these new discoveries to the knowledge base for all other animal species. It's hoped that the genome sequence will also provide common ground on which to unite studies of the chicken as a model species with those aimed at agriculturally-relevant applications. The current status of chicken genomics will be assessed with projections for its near and long term future.     

Building the sugarcane genome for biotechnology and identifying evolutionary trends

Background

Sugarcane is the source of sugar in all tropical and subtropical countries and is becoming increasingly important for bio-based fuels. However, its large (10 Gb), polyploid, complex genome has hindered genome based breeding efforts. Here we release the largest and most diverse set of sugarcane genome sequences to date, as part of an on-going initiative to provide a sugarcane genomic information resource, with the ultimate goal of producing a gold standard genome.

Results

Three hundred and seventeen chiefly euchromatic BACs were sequenced. A reference set of one thousand four hundred manually-annotated protein-coding genes was generated. A small RNA collection and a RNA-seq library were used to explore expression patterns and the sRNA landscape. In the sucrose and starch metabolism pathway, 16 non-redundant enzyme-encoding genes were identified. One of the sucrose pathway genes, sucrose-6-phosphate phosphohydrolase, is duplicated in sugarcane and sorghum, but not in rice and maize. A diversity analysis of the s6pp duplication region revealed haplotype-structured sequence composition. Examination of hom(e)ologous loci indicate both sequence structural and sRNA landscape variation. A synteny analysis shows that the sugarcane genome has expanded relative to the sorghum genome, largely due to the presence of transposable elements and uncharacterized intergenic and intronic sequences.

Conclusion

This release of sugarcane genomic sequences will advance our understanding of sugarcane genetics and contribute to the development of molecular tools for breeding purposes and gene discovery.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-540) contains supplementary material, which is available to authorized users.     

Effects of barley yellow mosaic disease resistant gene rym1 on the infection by strains of Barley yellow mosaic virus and Barley mild mosaic virus

Although a Chinese landrace of barley, Mokusekko 3, is completely resistant to all strains of Barley yellow mosaic virus (BaYMV) and Barley mild mosaic virus (BaMMV), and is known to have at least two resistant genes, rym1 and rym5, only rym5 has been utilized for BaYMV resistant barley breeding in Japan. In order to clarify the effect of rym1 on BaYMV and BaMMV, and to utilize the gene for resistant barley breeding, the susceptibilities of only rym1 carrying breeding lines against BaYMV and BaMMV were investigated. In the assessment of resistance to BaYMV-I, 341 F(2) populations derived from a cross between the resistant line Y4 with only rym1 and the susceptible cv Haruna Nijo shows that the segregation loosely fits a 1R:3S ratio (0.05 > P > 0.01), suggesting that the resistance is controlled by a single recessive gene, rym1. Further, none of the F(3) lines derived from the nine resistant F(2) plants showed any disease symptoms in the field infected by BaYMV-I. The same nine F(3) lines showed almost the same agronomic characters in the field infected by BaYMV-III as those in the uninfected field, apart from the symptom of showing numerous mosaics. This result indicates that the gene rym1 has an acceptable level of resistance to BaYMV-III. In the assessment of resistance to BaYMV-II, BaMMV-Ka1 and -Na1, an artificial infection method was adopted and the susceptibilities to those viruses were investigated. Although the control varieties, Ko A and Haruna Nijo, were infected with all of them, the rym1 gene carrying BC(2)F(3) lines were completely resistant to all strains. In summary, rym1 is completely resistant to BaYMV-I, -II, BaMMV-Ka1 and -Na1, and has an acceptable level of resistance to BaYMV-III. This study concludes with a discussion of the reason why the important resistance gene rym1 was eliminated along with resistant cultivars during breeding for resistance to BaYMV.     

Sequence Analysis of the Genome of Carnation (Dianthus caryophyllus L.)

The whole-genome sequence of carnation (Dianthus caryophyllus L.) cv. ‘Francesco’ was determined using a combination of different new-generation multiplex sequencing platforms. The total length of the non-redundant sequences was 568 887 315 bp, consisting of 45 088 scaffolds, which covered 91% of the 622 Mb carnation genome estimated by k-mer analysis. The N50 values of contigs and scaffolds were 16 644 bp and 60 737 bp, respectively, and the longest scaffold was 1 287 144 bp. The average GC content of the contig sequences was 36%. A total of 1050, 13, 92 and 143 genes for tRNAs, rRNAs, snoRNA and miRNA, respectively, were identified in the assembled genomic sequences. For protein-encoding genes, 43 266 complete and partial gene structures excluding those in transposable elements were deduced. Gene coverage was ∼98%, as deduced from the coverage of the core eukaryotic genes. Intensive characterization of the assigned carnation genes and comparison with those of other plant species revealed characteristic features of the carnation genome. The results of this study will serve as a valuable resource for fundamental and applied research of carnation, especially for breeding new carnation varieties. Further information on the genomic sequences is available at http://carnation.kazusa.or.jp.     

Distribution and characterization of simple sequence repeats in Gossypium raimondii genome

Simple sequence repeats (SSRs) can be derived from the complete genome sequence. These markers are important for gene mapping as well as marker-assisted selection (MAS). To develop SSRs for cotton gene mapping, we selected the complete genome sequence of Gossypium raimondii, which consisted of 4447 non-redundant scaffolds. Out of 775.2 Mb sequence examined, a total of 136,345 microsatellites were identified with a density of 5.69 kb per SSR in the G. raimondii genome leading to development of 112,177 primer pairs. The distributions of SSRs in the genome were non-random. Among the different motifs ranging from 1 to 6 bp, penta-nucleotide repeats were most abundant (30.5%), followed by tetra-nucleotide repeats (18.2%) and di-nucleotide repeats (16.9%). Among all identified 457 motif types, the most frequently occurring repeat motifs were poly-AT/TA, which accounted for 79.8% of the total di-nt SSRs, followed by AAAT/TTTA with 51.5% of the total tetra-nucleotede. Further, 18,834 microsatellites were detected from the protein-coding genes, and the frequency of gene containing SSRs was 46.0% in 40,976 genes of G. raimondii. These genome-based SSRs developed in the present study will lay the groundwork for developing large numbers of SSR markers for genetic mapping, gene discovery, genetic diversity analysis, and MAS breeding in cotton.     

Whole genome sequences are required to fully resolve the linkage disequilibrium structure of human populations

Background

An understanding of linkage disequilibrium (LD) structures in the human genome underpins much of medical genetics and provides a basis for disease gene mapping and investigating biological mechanisms such as recombination and selection. Whole genome sequencing (WGS) provides the opportunity to determine LD structures at maximal resolution.

Results

We compare LD maps constructed from WGS data with LD maps produced from the array-based HapMap dataset, for representative European and African populations. WGS provides up to 5.7-fold greater SNP density than array-based data and achieves much greater resolution of LD structure, allowing for identification of up to 2.8-fold more regions of intense recombination. The absence of ascertainment bias in variant genotyping improves the population representativeness of the WGS maps, and highlights the extent of uncaptured variation using array genotyping methodologies. The complete capture of LD patterns using WGS allows for higher genome-wide association study (GWAS) power compared to array-based GWAS, with WGS also allowing for the analysis of rare variation. The impact of marker ascertainment issues in arrays has been greatest for Sub-Saharan African populations where larger sample sizes and substantially higher marker densities are required to fully resolve the LD structure.

Conclusions

WGS provides the best possible resource for LD mapping due to the maximal marker density and lack of ascertainment bias. WGS LD maps provide a rich resource for medical and population genetics studies. The increasing availability of WGS data for large populations will allow for improved research utilising LD, such as GWAS and recombination biology studies.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1854-0) contains supplementary material, which is available to authorized users.     

New computational tools for brassica genome research

With the increasing quantities of Brassica genomic data being entered into the public domain and in preparation for the complete Brassica genome sequencing effort, there is a growing requirement for the structuring and detailed bioinformatic analysis of Brassica genomic information within a user-friendly database. At the Plant Biotechnology Centre, Melbourne, Australia, we have developed a series of tools and computational pipelines to assist in the processing and structuring of genomic data, to aid its application to agricultural biotechnology research. These tools include a sequence database, ASTRA, a sequence processing pipeline incorporating annotation against GenBank, SwissProt and Arabidopsis Gene Ontology (GO) data and tools for molecular marker discovery and comparative genome analysis. All sequences are mined for simple sequence repeat (SSR) molecular markers using 'SSR primer' and mapped onto the complete Arabidopsis thaliana genome by sequence comparison. The database may be queried using a text-based search of sequence annotation or GO terms, BLAST comparison against resident sequences, or by the position of candidate orthologues within the Arabidopsis genome. Tools have also been developed and applied to the discovery of single nucleotide polymorphism (SNP) molecular markers and the in silico mapping of Brassica BAC end sequences onto the Arabidopsis genome. Planned extensions to this resource include the integration of gene expression data and the development of an EnsEMBL-based genome viewer.