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Remnants of the Legume Ancestral Genome Preserved in Gene-Rich Regions: Insights from Lupinus angustifolius Physical, Genetic, and Comparative Mapping.

ABSTRACT: The narrow-leafed lupin (Lupinus angustifolius) was recently considered as a legume reference species. Genetic resources have been developed, including a draft genome sequence, linkage maps, nuclear DNA libraries, and cytogenetic chromosome-specific landmarks. Here, we used a complex approach, involving DNA fingerprinting, sequencing, genetic mapping, and molecular cytogenetics, to localize and analyze L. angustifolius gene-rich regions (GRRs). A L. angustifolius genomic bacterial artificial chromosome (BAC) library was screened with short sequence repeat (SSR)-based probes. Selected BACs were fingerprinted and assembled into contigs. BAC-end sequence (BES) annotation allowed us to choose clones for sequencing, targeting GRRs. Additionally, BESs were aligned to the scaffolds of the genome sequence. The genetic map was supplemented with 35 BES-derived markers, distributed in 14 linkage groups and tagging 37 scaffolds. The identified GRRs had an average gene density of 19.6 genes/100 kb and physical-to-genetic distance ratios of 11 to 109 kb/cM. Physical and genetic mapping was supported by multi-BAC-fluorescence in situ hybridization (FISH), and five new linkage groups were assigned to the chromosomes. Syntenic links to the genome sequences of five legume species (Medicago truncatula, Glycine max, Lotus japonicus, Phaseolus vulgaris, and Cajanus cajan) were identified. The comparative mapping of the two largest lupin GRRs provides novel evidence for ancient duplications in all of the studied species. These regions are conserved among representatives of the main clades of Papilionoideae. Furthermore, despite the complex evolution of legumes, some segments of the nuclear genome were not substantially modified and retained their quasi-ancestral structures. Cytogenetic markers anchored in these regions constitute a platform for heterologous mapping of legume genomes.

SUBMITTER: Ksiazkiewicz M

PROVIDER: S-EPMC4295026 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundThe narrow-leafed lupin, Lupinus angustifolius L., is a grain legume species with a relatively compact genome. The species has 2n = 40 chromosomes and its genome size is 960 Mbp/1C. During the last decade, L. angustifolius genomic studies have achieved several milestones, such as molecular-marker development, linkage maps, and bacterial artificial chromosome (BAC) libraries. Here, these resources were integratively used to identify and sequence two gene-rich regions (GRRs) of the genome.ResultsThe genome was screened with a probe representing the sequence of a microsatellite fragment length polymorphism (MFLP) marker linked to Phomopsis stem blight resistance. BAC clones selected by hybridization were subjected to restriction fingerprinting and contig assembly, and 232 BAC-ends were sequenced and annotated. BAC fluorescence in situ hybridization (BAC-FISH) identified eight single-locus clones. Based on physical mapping, cytogenetic localization, and BAC-end annotation, five clones were chosen for sequencing. Within the sequences of clones that hybridized in FISH to a single-locus, two large GRRs were identified. The GRRs showed strong and conserved synteny to Glycine max duplicated genome regions, illustrated by both identical gene order and parallel orientation. In contrast, in the clones with dispersed FISH signals, more than one-third of sequences were transposable elements. Sequenced, single-locus clones were used to develop 12 genetic markers, increasing the number of L. angustifolius chromosomes linked to appropriate linkage groups by five pairs.ConclusionsIn general, probes originating from MFLP sequences can assist genome screening and gene discovery. However, such probes are not useful for positional cloning, because they tend to hybridize to numerous loci. GRRs identified in L. angustifolius contained a low number of interspersed repeats and had a high level of synteny to the genome of the model legume G. max. Our results showed that not only was the gene nucleotide sequence conserved between soybean and lupin GRRs, but the order and orientation of particular genes in syntenic blocks was homologous, as well. These findings will be valuable to the forthcoming sequencing of the lupin genome.

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Remnants of the Legume Ancestral Genome Preserved in Gene-Rich Regions: Insights from Lupinus angustifolius Physical, Genetic, and Comparative Mapping.

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