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A BAC clone fingerprinting approach to the detection of human genome rearrangements
Martin Krzywinski, Ian Bosdet, Carrie Mathewson, Natasja Wye, Jay Brebner, Readman Chiu, Richard Corbett, Matthew Field, Darlene Lee, Trevor Pugh, Stas Volik, Asim Siddiqui, Steven Jones, Jacquie Schein, Collin Collins, Marco Marra
Genome Biology , 2007, DOI: 10.1186/gb-2007-8-10-r224
Abstract: The phenomenon of genomic heterogeneity, and the implications of this heterogeneity to human phenotypic diversity and disease, have recently been widely recognized [1-5], energizing efforts to develop catalogues of genomic variation [6-12]. Among efforts to understand the role and effect of genomic variability, landmark studies have described changes in the genetic landscape of both normal and diseased genomes [13-15], the presence of heterogeneity at different length scales [5,16] and variability within normal individuals of various ethnicities [17-19]. Genome rearrangements have been repeatedly linked to a variety of diseases, such as cancer [20] and mental retardation [21], and the evolution of alterations during disease progression continues to be an emphasis of current studies.Presently, various array-based methods, such as the 32 K bacterial artificial chromosome (BAC) array and Affy 100 K SNP array [21-23], are the most common approaches to detecting and localizing copy number variants, which are one class of genomic variation. The ubiquity of arrays is largely due to the fact that array experiments are relatively inexpensive, and collect information genome-wide. The advent of high-density oligonucleotide arrays, with probes spaced approximately every 5 kb, has increased the resolution of array methods to about 20-30 kb (multiple adjacent probes must confirm an aberration to be statistically significant) [21]. Despite their advantages, commonly available array-based methods have several shortcomings. These include the inability to: detect copy number neutral variants, such as balanced rearrangements; precisely delineate breakpoints and other fine structure details of genomic rearrangements; and directly provide substrates for functional sequence-based characterization once a rearrangement has been detected.Clone-based approaches have been developed to study genome structure, in part motivated by shortcomings of array-based methods [16,24,25]. In addition to t
Comparative analysis of pepper and tomato reveals euchromatin expansion of pepper genome caused by differential accumulation of Ty3/Gypsy-like elements
Minkyu Park, SungHwan Jo, Jin-Kyung Kwon, Jongsun Park, Jong Hwa Ahn, Seungill Kim, Yong-Hwan Lee, Tae-Jin Yang, Cheol-Goo Hur, Byoung-Cheorl Kang, Byung-Dong Kim, Doil Choi
BMC Genomics , 2011, DOI: 10.1186/1471-2164-12-85
Abstract: For sequence-level analysis, we generated 35.6 Mb of pepper genomic sequences from euchromatin enriched 1,245 pepper BAC clones. The comparative analysis of orthologous gene-rich regions between both species revealed insertion of transposons exclusively in the pepper sequences, maintaining the gene order and content. The most common type of the transposon found was the LTR retrotransposon. Phylogenetic comparison of the LTR retrotransposons revealed that two groups of Ty3/Gypsy-like elements (Tat and Athila) were overly accumulated in the pepper genome. The FISH analysis of the pepper Tat elements showed a random distribution in heterochromatic and euchromatic regions, whereas the tomato Tat elements showed heterochromatin-preferential accumulation.Compared to tomato pepper euchromatin doubled its size by differential accumulation of a specific group of Ty3/Gypsy-like elements. Our results could provide an insight on the mechanism of genome evolution in the Solanaceae family.The Solanaceae is an unusually divergent family consisting of approximately 90 genera and 3,000-4,000 species [1]. Members of the Solanaceae have evolved into extremely divergent forms, ranging from trees to annual herbs, and they occupy diverse habitats ranging from deserts to aquatic areas [1]. Such hyper-diversity in one family makes it useful to study plant adaptation and diversification. Despite this diversity, all Solanaceous species evolved during the last 40 million years [2]. Furthermore, almost all members share the same chromosome number (x = 12) [2].To date, diversity within the Solanaceae has been studied by comparative genome analyses using common genetic markers. As a result, we know that the Solanaceae genomes have undergone relatively small numbers of chromosomal rearrangements (e.g., about 5 rearrangements between potato and tomato and about 30 rearrangements between pepper and tomato), maintaining well-conserved gene content and order [3-8]. The conservation of the Solanaceae
Comparative analysis of catfish BAC end sequences with the zebrafish genome
Hong Liu, Yanliang Jiang, Shaolin Wang, Parichart Ninwichian, Benjaporn Somridhivej, Peng Xu, Jason Abernathy, Huseyin Kucuktas, Zhanjiang Liu
BMC Genomics , 2009, DOI: 10.1186/1471-2164-10-592
Abstract: We reported the generation of 43,000 BAC end sequences and their applications for comparative genome analysis in catfish. Using these and the additional 20,000 existing BAC end sequences as a resource along with linkage mapping and existing physical map, conserved syntenic regions were identified between the catfish and zebrafish genomes. A total of 10,943 catfish BAC end sequences (17.3%) had significant BLAST hits to the zebrafish genome (cutoff value ≤ e-5), of which 3,221 were unique gene hits, providing a platform for comparative mapping based on locations of these genes in catfish and zebrafish. Genetic linkage mapping of microsatellites associated with contigs allowed identification of large conserved genomic segments and construction of super scaffolds.BAC end sequences and their associated polymorphic markers are great resources for comparative genome analysis in catfish. Highly conserved chromosomal regions were identified to exist between catfish and zebrafish. However, it appears that the level of conservation at local genomic regions are high while a high level of chromosomal shuffling and rearrangements exist between catfish and zebrafish genomes. Orthologous regions established through comparative analysis should facilitate both structural and functional genome analysis in catfish.Comparative mapping is a powerful tool to transfer genomic information from sequenced genomes to closely related species for which whole genome sequence data are not yet available. Such an approach was initially demonstrated by Fujiyama et al. [1] for the construction of the human-chimpanzee comparative map. In these closely related primate species, approximately 98% of chimpanzee BAC end sequences (BES) had significant BLAST hits to the human genome sequence allowing putative orthologues to be identified [1]. A similar approach was used for the construction of the human-mouse comparative map [2]. Subsequently, this approach was extensively used in mammals including construc
Genome Mapping and Molecular Breeding of Tomato  [PDF]
Majid R. Foolad
International Journal of Plant Genomics , 2007, DOI: 10.1155/2007/64358
Abstract: The cultivated tomato, Lycopersicon esculentum, is the second most consumed vegetable worldwide and a well-studied crop species in terms of genetics, genomics, and breeding. It is one of the earliest crop plants for which a genetic linkage map was constructed, and currently there are several molecular maps based on crosses between the cultivated and various wild species of tomato. The high-density molecular map, developed based on an L. esculentum×L. pennellii cross, includes more than 2200 markers with an average marker distance of less than 1 cM and an average of 750 kbp per cM. Different types of molecular markers such as RFLPs, AFLPs, SSRs, CAPS, RGAs, ESTs, and COSs have been developed and mapped onto the 12 tomato chromosomes. Markers have been used extensively for identification and mapping of genes and QTLs for many biologically and agriculturally important traits and occasionally for germplasm screening, fingerprinting, and marker-assisted breeding. The utility of MAS in tomato breeding has been restricted largely due to limited marker polymorphism within the cultivated species and economical reasons. Also, when used, MAS has been employed mainly for improving simply-inherited traits and not much for improving complex traits. The latter has been due to unavailability of reliable PCR-based markers and problems with linkage drag. Efforts are being made to develop high-throughput markers with greater resolution, including SNPs. The expanding tomato EST database, which currently includes ∼214 000 sequences, the new microarray DNA chips, and the ongoing sequencing project are expected to aid development of more practical markers. Several BAC libraries have been developed that facilitate map-based cloning of genes and QTLs. Sequencing of the euchromatic portions of the tomato genome is paving the way for comparative and functional analysis of important genes and QTLs.
The European sea bass Dicentrarchus labrax genome puzzle: comparative BAC-mapping and low coverage shotgun sequencing
Heiner Kuhl, Alfred Beck, Grzegorz Wozniak, Adelino VM Canario, Filip AM Volckaert, Richard Reinhardt
BMC Genomics , 2010, DOI: 10.1186/1471-2164-11-68
Abstract: End sequencing of a sea bass genomic BAC-library enabled the comparative mapping of the sea bass genome using the three-spined stickleback Gasterosteus aculeatus genome as a reference. BAC-end sequences (102,690) were aligned to the stickleback genome. The number of mappable BACs was improved using a two-fold coverage WGS dataset of sea bass resulting in a comparative BAC-map covering 87% of stickleback chromosomes with 588 BAC-contigs. The minimum size of 83 contigs covering 50% of the reference was 1.2 Mbp; the largest BAC-contig comprised 8.86 Mbp. More than 22,000 BAC-clones aligned with both ends to the reference genome. Intra-chromosomal rearrangements between sea bass and stickleback were identified. Size distributions of mapped BACs were used to calculate that the genome of sea bass may be only 1.3 fold larger than the 460 Mbp stickleback genome.The BAC map is used for sequencing single BACs or BAC-pools covering defined genomic entities by second generation sequencing technologies. Together with the WGS dataset it initiates a sea bass genome sequencing project. This will allow the quantification of polymorphisms through resequencing, which is important for selecting highly performing domesticated fish.Teleost fishes are the most diverse group of vertebrates, with approximately 28,000 species, which have colonized a range of aquatic environments and display a variety of biochemical, physiological and morphological adaptations [1,2]. Because of this diversity and their position at the base of the vertebrate phylogeny, some species are considered good models of evolution, development and human diseases [3-6]. For this reason, teleost species were among the first vertebrate genomes to be sequenced: the green spotted pufferfish, Tetraodon nigroviridis [7]and the fugu Takifugu rubripes [8] for their relatively small compact genome; the medaka Oryzias latipes [9] and the zebrafish Danio rerio [10] for their value as developmental models, short life cycle, ease of
Large-scale analysis of full-length cDNAs from the tomato (Solanum lycopersicum) cultivar Micro-Tom, a reference system for the Solanaceae genomics
Koh Aoki, Kentaro Yano, Ayako Suzuki, Shingo Kawamura, Nozomu Sakurai, Kunihiro Suda, Atsushi Kurabayashi, Tatsuya Suzuki, Taneaki Tsugane, Manabu Watanabe, Kazuhide Ooga, Maiko Torii, Takanori Narita, Tadasu Shin-i, Yuji Kohara, Naoki Yamamoto, Hideki Takahashi, Yuichiro Watanabe, Mayumi Egusa, Motoichiro Kodama, Yuki Ichinose, Mari Kikuchi, Sumire Fukushima, Akiko Okabe, Tsutomu Arie, Yuko Sato, Katsumi Yazawa, Shinobu Satoh, Toshikazu Omura, Hiroshi Ezura, Daisuke Shibata
BMC Genomics , 2010, DOI: 10.1186/1471-2164-11-210
Abstract: To accelerate the progress in tomato genomics, we developed a collection of fully-sequenced 13,227 Micro-Tom full-length cDNAs. By checking redundant sequences, coding sequences, and chimeric sequences, a set of 11,502 non-redundant full-length cDNAs (nrFLcDNAs) was generated. Analysis of untranslated regions demonstrated that tomato has longer 5'- and 3'-untranslated regions than most other plants but rice. Classification of functions of proteins predicted from the coding sequences demonstrated that nrFLcDNAs covered a broad range of functions. A comparison of nrFLcDNAs with genes of sixteen plants facilitated the identification of tomato genes that are not found in other plants, most of which did not have known protein domains. Mapping of the nrFLcDNAs onto currently available tomato genome sequences facilitated prediction of exon-intron structure. Introns of tomato genes were longer than those of Arabidopsis and rice. According to a comparison of exon sequences between the nrFLcDNAs and the tomato genome sequences, the frequency of nucleotide mismatch in exons between Micro-Tom and the genome-sequencing cultivar (Heinz 1706) was estimated to be 0.061%.The collection of Micro-Tom nrFLcDNAs generated in this study will serve as a valuable genomic tool for plant biologists to bridge the gap between basic and applied studies. The nrFLcDNA sequences will help annotation of the tomato whole-genome sequence and aid in tomato functional genomics and molecular breeding. Full-length cDNA sequences and their annotations are provided in the database KaFTom http://www.pgb.kazusa.or.jp/kaftom/ webcite via the website of the National Bioresource Project Tomato http://tomato.nbrp.jp webcite.The Solanaceae family comprises 1000-2000 species that show wide morphological variability and ecological adaptability [1]. This taxon includes a number of vegetable crops including fruit-bearing vegetables, tuber-bearing vegetables, and ornamental plants, many of which have economic importan
Convergence of developmental mutants into a single tomato model system: 'Micro-Tom' as an effective toolkit for plant development research
Rogério F Carvalho, Marcelo L Campos, Lilian E Pino, Simone L Crestana, Agustin Zs?g?n, Joni E Lima, Vagner A Benedito, Lázaro EP Peres
Plant Methods , 2011, DOI: 10.1186/1746-4811-7-18
Abstract: We took advantage of the small size and rapid life cycle of the tomato cultivar Micro-Tom (MT) to create near-isogenic lines (NILs) by introgressing a suite of hormonal and photomorphogenetic mutations (altered sensitivity or endogenous levels of auxin, ethylene, abscisic acid, gibberellin, brassinosteroid, and light response) into this genetic background. To demonstrate the usefulness of this collection, we compared developmental traits between the produced NILs. All expected mutant phenotypes were expressed in the NILs. We also created NILs harboring the wild type alleles for dwarf, self-pruning and uniform fruit, which are mutations characteristic of MT. This amplified both the applications of the mutant collection presented here and of MT as a genetic model system.The community resource presented here is a useful toolkit for plant research, particularly for future studies in plant development, which will require the simultaneous observation of the effect of various hormones, signaling pathways and crosstalk.In addition to its worldwide cultivation and economic importance, tomato (Solanum lycopersicum L.) has several characteristics that make it a convenient model plant species, such as a relatively compact genome (950 Mb) combined with a marker-saturated genetic linkage map [1], rich germplasm collections (Tomato Genetics Resource Center) [2] and highly efficient transformation protocols [3]. The pre-released annotated genome sequence [1] appears set to establish tomato as a prominent model system for research into plant genetics and physiology. The tomato is poised to become an alternative model plant to Arabidopsis thaliana due to its diverse developmental traits not found in Arabidopsis. These traits include: the photoperiod-independent sympodial flowering and the formation of fleshy climacteric fruits, compound leaves, mycorrhizal roots and glandular trichomes.The convenient small size and amenability to large-scale cultivation of Arabidopsis are also found
A BAC end view of the Musa acuminata genome
Foo Cheung, Christopher D Town
BMC Plant Biology , 2007, DOI: 10.1186/1471-2229-7-29
Abstract: BAC end sequencing generated 6,252 reads representing 4,420,944 bp, including 2,979 clone pairs with an average read length after cleaning and filtering of 707 bp. All sequences have been submitted to GenBank, with the accession numbers DX451975 – DX458350. The BAC end-sequences, were searched against several databases and significant homology was found to mitochondria and chloroplast (2.6%), transposons and repetitive sequences (36%) and proteins (11%). Functional interpretation of the protein matches was carried out by Gene Ontology assignments from matches to Arabidopsis and was shown to cover a broad range of categories. From protein matching regions of Musa BAC end-sequences, it was determined that the GC content of coding regions was 47%. Where protein matches encompassed a start codon, GC content as a function of position (5' to 3') across 129 bp sliding windows generates a "rice-like" gradient. A total of 352 potential SSR markers were discovered. The most abundant simple sequence repeats in four size categories were AT-rich. After filtering mitochondria and chloroplast matches, thousands of BAC end-sequences had a significant BLASTN match to the Oryza sativa and Arabidopsis genome sequence. Of these, a small number of BAC end-sequence pairs were shown to map to neighboring regions of the Oryza sativa genome representing regions of potential microsynteny.Database searches with the BAC end-sequences and ab initio analysis identified those reads likely to contain transposons, repeat sequences, proteins and simple sequence repeats. Approximately 600 BAC end-sequences contained protein sequences that were not found in the existing available Musa expressed sequence tags, repeat or transposon databases. In addition, gene statistics, GC content and profile could also be estimated based on the region matching the top protein hit. A small number of BAC end pair sequences can be mapped to neighboring regions of the Oryza sativa representing regions of potential micros
Ontology for Genome Comparison and Genomic Rearrangements  [PDF]
Keith Flanagan,Robert Stevens,Matthew Pocock,Pete Lee,Anil Wipat
Comparative and Functional Genomics , 2004, DOI: 10.1002/cfg.436
Abstract: We present an ontology for describing genomes, genome comparisons, their evolution and biological function. This ontology will support the development of novel genome comparison algorithms and aid the community in discussing genomic evolution. It provides a framework for communication about comparative genomics, and a basis upon which further automated analysis can be built. The nomenclature defined by the ontology will foster clearer communication between biologists, and also standardize terms used by data publishers in the results of analysis programs. The overriding aim of this ontology is the facilitation of consistent annotation of genomes through computational methods, rather than human annotators. To this end, the ontology includes definitions that support computer analysis and automated transfer of annotations between genomes, rather than relying upon human mediation.
Analysis of 90 Mb of the potato genome reveals conservation of gene structures and order with tomato but divergence in repetitive sequence composition
Wei Zhu, Shu Ouyang, Marina Iovene, Kimberly O'Brien, Hue Vuong, Jiming Jiang, C Robin Buell
BMC Genomics , 2008, DOI: 10.1186/1471-2164-9-286
Abstract: In this study, we report on analysis 89.9 Mb of potato genomic sequence representing 10.2% of the genome generated through end sequencing of a potato bacterial artificial chromosome (BAC) clone library (87 Mb) and sequencing of 22 potato BAC clones (2.9 Mb). The GC content of potato is very similar to Solanum lycopersicon (tomato) and other dicotyledonous species yet distinct from the monocotyledonous grass species, Oryza sativa. Parallel analyses of repetitive sequences in potato and tomato revealed substantial differences in their abundance, 34.2% in potato versus 46.3% in tomato, which is consistent with the increased genome size per haploid genome of these two Solanum species. Specific classes and types of repetitive sequences were also differentially represented between these two species including a telomeric-related repetitive sequence, ribosomal DNA, and a number of unclassified repetitive sequences. Comparative analyses between tomato and potato at the gene level revealed a high level of conservation of gene content, genic feature, and gene order although discordances in synteny were observed.Genomic level analyses of potato and tomato confirm that gene sequence and gene order are conserved between these solanaceous species and that this conservation can be leveraged in genomic applications including cross-species annotation and genome sequencing initiatives. While tomato and potato share genic features, they differ in their repetitive sequence content and composition suggesting that repetitive sequences may have a more significant role in shaping speciation than previously reported.The potato (Solanum tuberosum) tuber is a specialized underground storage organ that develops from modified stems termed stolons. Although the tuber is primarily composed of starch, it also contains high levels of proteins and due to its importance as a food source, a prime focus in potato research has been tuber quality [1-6]. Another key focus in potato research is disease resi
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