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The first generation of a BAC-based physical map of Brassica rapa.

ABSTRACT: BACKGROUND: The genus Brassica includes the most extensively cultivated vegetable crops worldwide. Investigation of the Brassica genome presents excellent challenges to study plant genome evolution and divergence of gene function associated with polyploidy and genome hybridization. A physical map of the B. rapa genome is a fundamental tool for analysis of Brassica "A" genome structure. Integration of a physical map with an existing genetic map by linking genetic markers and BAC clones in the sequencing pipeline provides a crucial resource for the ongoing genome sequencing effort and assembly of whole genome sequences. RESULTS: A genome-wide physical map of the B. rapa genome was constructed by the capillary electrophoresis-based fingerprinting of 67,468 Bacterial Artificial Chromosome (BAC) clones using the five restriction enzyme SNaPshot technique. The clones were assembled into contigs by means of FPC v8.5.3. After contig validation and manual editing, the resulting contig assembly consists of 1,428 contigs and is estimated to span 717 Mb in physical length. This map provides 242 anchored contigs on 10 linkage groups to be served as seed points from which to continue bidirectional chromosome extension for genome sequencing. CONCLUSION: The map reported here is the first physical map for Brassica "A" genome based on the High Information Content Fingerprinting (HICF) technique. This physical map will serve as a fundamental genomic resource for accelerating genome sequencing, assembly of BAC sequences, and comparative genomics between Brassica genomes. The current build of the B. rapa physical map is available at the B. rapa Genome Project website for the user community.

SUBMITTER: Mun JH

PROVIDER: S-EPMC2432078 | biostudies-literature | 2008

REPOSITORIES: biostudies-literature

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The first generation of a BAC-based physical map of Brassica rapa.

Mun Jeong-Hwan JH Kwon Soo-Jin SJ Yang Tae-Jin TJ Kim Hye-Sun HS Choi Beom-Soon BS Baek Seunghoon S Kim Jung Sun JS Jin Mina M Kim Jin A JA Lim Myung-Ho MH Lee Soo In SI Kim Ho-Il HI Kim Hyungtae H Lim Yong Pyo YP Park Beom-Seok BS

BMC genomics 20080612

<h4>Background</h4>The genus Brassica includes the most extensively cultivated vegetable crops worldwide. Investigation of the Brassica genome presents excellent challenges to study plant genome evolution and divergence of gene function associated with polyploidy and genome hybridization. A physical map of the B. rapa genome is a fundamental tool for analysis of Brassica "A" genome structure. Integration of a physical map with an existing genetic map by linking genetic markers and BAC clones in ...[more]

PMID: 18549474

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Project description:BackgroundRainbow trout (Oncorhynchus mykiss) are the most-widely cultivated cold freshwater fish in the world and an important model species for many research areas. Coupling great interest in this species as a research model with the need for genetic improvement of aquaculture production efficiency traits justifies the continued development of genomics research resources. Many quantitative trait loci (QTL) have been identified for production and life-history traits in rainbow trout. A bacterial artificial chromosome (BAC) physical map is needed to facilitate fine mapping of QTL and the selection of positional candidate genes for incorporation in marker-assisted selection (MAS) for improving rainbow trout aquaculture production. This resource will also facilitate efforts to obtain and assemble a whole-genome reference sequence for this species.ResultsThe physical map was constructed from DNA fingerprinting of 192,096 BAC clones using the 4-color high-information content fingerprinting (HICF) method. The clones were assembled into physical map contigs using the finger-printing contig (FPC) program. The map is composed of 4,173 contigs and 9,379 singletons. The total number of unique fingerprinting fragments (consensus bands) in contigs is 1,185,157, which corresponds to an estimated physical length of 2.0 Gb. The map assembly was validated by 1) comparison with probe hybridization results and agarose gel fingerprinting contigs; and 2) anchoring large contigs to the microsatellite-based genetic linkage map.ConclusionThe production and validation of the first BAC physical map of the rainbow trout genome is described in this paper. We are currently integrating this map with the NCCCWA genetic map using more than 200 microsatellites isolated from BAC end sequences and by identifying BACs that harbor more than 300 previously mapped markers. The availability of an integrated physical and genetic map will enable detailed comparative genome analyses, fine mapping of QTL, positional cloning, selection of positional candidate genes for economically important traits and the incorporation of MAS into rainbow trout breeding programs.

Project description:BACKGROUND: Half-smooth tongue sole (Cynoglossus semilaevis Günther) has been exploited as a commercially important cultured marine flatfish, and female grows 2-3 times faster than male. Genetic studies, especially on the chromosomal sex-determining system of this species, have been carried out in the last decade. Although the genome of half-smooth tongue sole was relatively small (626.9 Mb), there are still some difficulties in the high-quality assembly of the next generation genome sequencing reads without the assistance of a physical map, especially for the W chromosome of this fish due to abundance of repetitive sequences. The objective of this study is to construct a bacterial artificial chromosome (BAC)-based physical map for half-smooth tongue sole with the method of high information content fingerprinting (HICF). RESULTS: A physical map of half-smooth tongue sole was constructed with 30, 294 valid fingerprints (7.5 × genome coverage) with a tolerance of 4 and an initial cutoff of 1e-60. A total of 29,709 clones were assembled into 1,485 contigs with an average length of 539 kb and a N50 length of 664 kb. There were 394 contigs longer than the N50 length, and these contigs will be a useful resource for future integration with linkage map and whole genome sequence assembly. The estimated physical length of the assembled contigs was 797 Mb, representing approximately 1.27 coverage of the half-smooth tongue sole genome. The largest contig contained 410 BAC clones with a physical length of 3.48 Mb. Almost all of the 676 BAC clones (99.9%) in the 21 randomly selected contigs were positively validated by PCR assays, thereby confirming the reliability of the assembly. CONCLUSIONS: A first generation BAC-based physical map of half-smooth tongue sole was constructed with high reliability. The map will promote genetic improvement programs of this fish, especially integration of physical and genetic maps, fine-mappings of important gene and/or QTL, comparative and evolutionary genomics studies, as well as whole genome sequence assembly.

Project description:BackgroundCucumis melo (melon) belongs to the Cucurbitaceae family, whose economic importance among horticulture crops is second only to Solanaceae. Melon has high intra-specific genetic variation, morphologic diversity and a small genome size (450 Mb), which make this species suitable for a great variety of molecular and genetic studies that can lead to the development of tools for breeding varieties of the species. A number of genetic and genomic resources have already been developed, such as several genetic maps and BAC genomic libraries. These tools are essential for the construction of a physical map, a valuable resource for map-based cloning, comparative genomics and assembly of whole genome sequencing data. However, no physical map of any Cucurbitaceae has yet been developed. A project has recently been started to sequence the complete melon genome following a whole-genome shotgun strategy, which makes use of massive sequencing data. A BAC-based melon physical map will be a useful tool to help assemble and refine the draft genome data that is being produced.ResultsA melon physical map was constructed using a 5.7 x BAC library and a genetic map previously developed in our laboratories. High-information-content fingerprinting (HICF) was carried out on 23,040 BAC clones, digesting with five restriction enzymes and SNaPshot labeling, followed by contig assembly with FPC software. The physical map has 1,355 contigs and 441 singletons, with an estimated physical length of 407 Mb (0.9 x coverage of the genome) and the longest contig being 3.2 Mb. The anchoring of 845 BAC clones to 178 genetic markers (100 RFLPs, 76 SNPs and 2 SSRs) also allowed the genetic positioning of 183 physical map contigs/singletons, representing 55 Mb (12%) of the melon genome, to individual chromosomal loci. The melon FPC database is available for download at http://melonomics.upv.es/static/files/public/physical_map/.ConclusionsHere we report the construction of the first physical map of a Cucurbitaceae species described so far. The physical map was integrated with the genetic map so that a number of physical contigs, representing 12% of the melon genome, could be anchored to known genetic positions. The data presented is already helping to improve the quality of the melon genomic sequence available as a result of a project currently being carried out in Spain, adopting a whole genome shotgun approach based on 454 sequencing data.

Project description:BackgroundIn view of the immense value of Brassica rapa in the fields of agriculture and molecular biology, the multinational Brassica rapa Genome Sequencing Project (BrGSP) was launched in 2003 by five countries. The developing BrGSP has valuable resources for the community, including a reference genetic map and seed BAC sequences. Although the initial B. rapa linkage map served as a reference for the BrGSP, there was ambiguity in reconciling the linkage groups with the ten chromosomes of B. rapa. Consequently, the BrGSP assigned each of the linkage groups to the project members as chromosome substitutes for sequencing.ResultsWe identified simple sequence repeat (SSR) motifs in the B. rapa genome with the sequences of seed BACs used for the BrGSP. By testing 749 amplicons containing SSR motifs, we identified polymorphisms that enabled the anchoring of 188 BACs onto the B. rapa reference linkage map consisting of 719 loci in the 10 linkage groups with an average distance of 1.6 cM between adjacent loci. The anchored BAC sequences enabled the identification of 30 blocks of conserved synteny, totaling 534.9 cM in length, between the genomes of B. rapa and Arabidopsis thaliana. Most of these were consistent with previously reported duplication and rearrangement events that differentiate these genomes. However, we were able to identify the collinear regions for seven additional previously uncharacterized sections of the A genome. Integration of the linkage map with the B. rapa cytogenetic map was accomplished by FISH with probes representing 20 BAC clones, along with probes for rDNA and centromeric repeat sequences. This integration enabled unambiguous alignment and orientation of the maps representing the 10 B. rapa chromosomes.ConclusionWe developed a second generation reference linkage map for B. rapa, which was aligned unambiguously to the B. rapa cytogenetic map. Furthermore, using our data, we confirmed and extended the comparative genome analysis between B. rapa and A. thaliana. This work will serve as a basis for integrating the genetic, physical, and chromosome maps of the BrGSP, as well as for studies on polyploidization, speciation, and genome duplication in the genus Brassica.

Project description:BACKGROUND: The genus Aquilegia, consisting of approximately 70 taxa, is a member of the basal eudicot lineage, Ranuculales, which is evolutionarily intermediate between monocots and core eudicots, and represents a relatively unstudied clade in the angiosperm phylogenetic tree that bridges the gap between these two major plant groups. Aquilegia species are closely related and their distribution covers highly diverse habitats. These provide rich resources to better understand the genetic basis of adaptation to different pollinators and habitats that in turn leads to rapid speciation. To gain insights into the genome structure and facilitate gene identification, comparative genomics and whole-genome shotgun sequencing assembly, BAC-based genomics resources are of crucial importance. RESULTS: BAC-based genomic resources, including two BAC libraries, a physical map with anchored markers and BAC end sequences, were established from A. formosa. The physical map was composed of a total of 50,155 BAC clones in 832 contigs and 3939 singletons, covering 21X genome equivalents. These contigs spanned a physical length of 689.8 Mb (~2.3X of the genome) suggesting the complex heterozygosity of the genome. A set of 197 markers was developed from ESTs induced by drought-stress, or involved in anthocyanin biosynthesis or floral development, and was integrated into the physical map. Among these were 87 genetically mapped markers that anchored 54 contigs, spanning 76.4 Mb (25.5%) across the genome. Analysis of a selection of 12,086 BAC end sequences (BESs) from the minimal tiling path (MTP) allowed a preview of the Aquilegia genome organization, including identification of transposable elements, simple sequence repeats and gene content. Common repetitive elements previously reported in both monocots and core eudicots were identified in Aquilegia suggesting the value of this genome in connecting the two major plant clades. Comparison with sequenced plant genomes indicated a higher similarity to grapevine (Vitis vinifera) than to rice and Arabidopsis in the transcriptomes. CONCLUSIONS: The A. formosa BAC-based genomic resources provide valuable tools to study Aquilegia genome. Further integration of other existing genomics resources, such as ESTs, into the physical map should enable better understanding of the molecular mechanisms underlying adaptive radiation and elaboration of floral morphology.

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The first generation of a BAC-based physical map of Brassica rapa.

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The first generation of a BAC-based physical map of Brassica rapa.

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