Role of Fluorescence in situ Hybridization in Sequencing the Tomato Genome

The tomato (Solanum lycopersicum L.) genome is being sequenced by a consortium of laboratories in 10 countries. Seventy-seven percent of the tomato genome (DNA) is located in repeat-rich, gene-poor, pericentric heterochromatin, while 23% of the genome is located in repeat-poor, gene-rich, distal euchromatin. It is estimated that approximately 90% of tomato’s nuclear genes can be characterized by limiting the sequencing effort to euchromatin while avoiding the problems involved in sequencing the repetitive DNA in heterochromatin. Sequencing is being performed on tomato nuclear DNA cloned into bacterial artificial chromosome (BAC) vectors. Fluorescence in situ hybridization (FISH) is used to help direct the sequencing effort by cytologically demonstrating the location of selected BACs on tomato chromosomes. While mitotic metaphase chromosomes are too short and compact for this purpose, long pachytene chromosomes are ideal. BACs localized in euchromatin can be used confidently as anchors for the assembly of BAC contigs that extend through the euchromatic length of each chromosome arm. Another important role for FISH is identification of BACs near telomeres and near borders with pericentric heterochromatin to indicate that sequencing should not extend much further. This role of FISH is enhanced by our ability to estimate base pair distances between localized BACs and these chromosomal features. Finally, it is noteworthy that when BAC-FISH is combined with chromosomal in situ suppression (CISS) hybridization to block repeats and localize single/low copy sequences, the great majority of BACs localize to single sites. This observation is consistent with tomato being an ancient diploid.

[1]  L. Anderson,et al.  Electron microscopic immunogold localization of recombination-related proteins in spreads of synaptonemal complexes from tomato microsporocytes. , 2009, Methods in molecular biology.

[2]  Yuling Bai,et al.  High-resolution chromosome mapping of BACs using multi-colour FISH and pooled-BAC FISH as a backbone for sequencing tomato chromosome 6. , 2008, The Plant journal : for cell and molecular biology.

[3]  E. Stockinger,et al.  A Retrotransposon-Mediated Gene Duplication Underlies Morphological Variation of Tomato Fruit , 2008, Science.

[4]  D. Peterson,et al.  Isolation of milligram quantities of nuclear DNA from tomato (Lycopersicon esculentum), A plant containing high levels of polyphenolic compounds , 1997, Plant Molecular Biology Reporter.

[5]  L. Anderson,et al.  Predicting and Testing Physical Locations of Genetically Mapped Loci on Tomato Pachytene Chromosome 1 , 2007, Genetics.

[6]  D. Ashlock,et al.  Nearly Identical Paralogs: Implications for Maize (Zea mays L.) Genome Evolution , 2007, Genetics.

[7]  X. Zhong,et al.  High-resolution mapping on pachytene chromosomes and extended DNA fibres by fluorescencein-situ hybridisation , 1996, Plant Molecular Biology Reporter.

[8]  E. D. Earle,et al.  Nuclear DNA content of some important plant species , 1991, Plant Molecular Biology Reporter.

[9]  S. Tanksley,et al.  Combining Bioinformatics and Phylogenetics to Identify Large Sets of Single-Copy Orthologous Genes (COSII) for Comparative, Evolutionary and Systematic Studies: A Test Case in the Euasterid Plant Clade , 2006, Genetics.

[10]  S. Tanksley,et al.  Euchromatin and Pericentromeric Heterochromatin: Comparative Composition in the Tomato Genome , 2006, Genetics.

[11]  Carolyn J Lawrence,et al.  Predicting Chromosomal Locations of Genetically Mapped Loci in Maize Using the Morgan2McClintock Translator , 2006, Genetics.

[12]  Dawei Li,et al.  The Genomes of Oryza sativa: A History of Duplications , 2005, PLoS biology.

[13]  G. H. Jones,et al.  A light microscopic atlas of meiosis inArabidopsis thaliana , 1996, Chromosome Research.

[14]  D. Ward,et al.  Delineation of individual human chromosomes in metaphase and interphase cells by in situ suppression hybridization using recombinant DNA libraries , 1988, Human Genetics.

[15]  G. Khush,et al.  Studies on the linkage map of chromosome 4 of the tomato and on the transmission of induced deficiencies , 1967, Genetica.

[16]  M. Ramanna,et al.  Structure of and homology between pachytene and somatic metaphase chromosomes of the tomato , 1967, Genetica.

[17]  Jessica A Schlueter,et al.  Mining EST databases to resolve evolutionary events in major crop species. , 2004, Genome.

[18]  S. Jackson,et al.  Segmental duplications within the Glycine max genome revealed by fluorescence in situ hybridization of bacterial artificial chromosomes. , 2004, Genome.

[19]  Guillaume Blanc,et al.  Widespread Paleopolyploidy in Model Plant Species Inferred from Age Distributions of Duplicate Genes , 2004, The Plant Cell Online.

[20]  Beat Keller,et al.  Ancestral genome duplication in rice. , 2004, Genome.

[21]  M. Ploeg,et al.  Use of whole cosmid cloned genomic sequences for chromosomal localization by non-radioactive in situ hybridization , 1987, Human Genetics.

[22]  S. Chang.  Cytogenetic and molecular studies on tomato chromosomes using diploid tomato and tomato monosomic additions in tetraploid potato , 2004 .

[23]  L. Anderson,et al.  A model for chromosome structure during the mitotic and meiotic cell cycles , 2004, Chromosome Research.

[24]  R. Wing,et al.  Resolution of fluorescence in-situ hybridization mapping on rice mitotic prometaphase chromosomes, meiotic pachytene chromosomes and extended DNA fibers , 2004, Chromosome Research.

[25]  Jonathan F. Wendel,et al.  Genome evolution in polyploids , 2004, Plant Molecular Biology.

[26]  Ilia J. Leitch,et al.  Comparisons with Caenorhabditis (∼100 Mb) and Drosophila (∼175 Mb) Using Flow Cytometry Show Genome Size in Arabidopsis to be ∼157 Mb and thus ∼25 % Larger than the Arabidopsis Genome Initiative Estimate of ∼125 Mb , 2003 .

[27]  R. Wing,et al.  Toward a cytological characterization of the rice genome. , 2001, Genome research.

[28]  T. Bisseling,et al.  Integration of the FISH pachytene and genetic maps of Medicago truncatula. , 2001, The Plant journal : for cell and molecular biology.

[29]  D. G. Brown,et al.  The origins of genomic duplications in Arabidopsis. , 2000, Science.

[30]  S. Tanksley,et al.  Comparing sequenced segments of the tomato and Arabidopsis genomes: large-scale duplication followed by selective gene loss creates a network of synteny. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[31]  T. Wood,et al.  A deep-coverage tomato BAC library and prospects toward development of an STC framework for genome sequencing. , 2000, Genome research.

[32]  J. Hans de Jong,et al.  High resolution FISH in plants - techniques and applications. , 1999, Trends in plant science.

[33]  D. Peterson,et al.  Localization of single- and low-copy sequences on tomato synaptonemal complex spreads using fluorescence in situ hybridization (FISH). , 1999, Genetics.

[34]  J. Bodeau,et al.  FISH to meiotic pachytene chromosomes of tomato locates the root-knot nematode resistance gene Mi-1 and the acid phosphatase gene Aps-1 near the junction of euchromatin and pericentromeric heterochromatin of chromosome arms 6S and 6L, respectively , 1999, Theoretical and Applied Genetics.

[35]  W. Pearson,et al.  Characterization of the tomato (Lycopersicon esculentum) genome using in vitro and in situ DNA reassociation , 1998 .

[36]  S. Armstrong,et al.  Cytogenetics for the model system Arabidopsis thaliana. , 1998, The Plant journal : for cell and molecular biology.

[37]  B. Gaut,et al.  DNA sequence evidence for the segmental allotetraploid origin of maize. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[38]  R. Wing,et al.  A rapid procedure for the isolation of C0t-1 DNA from plants. , 1997, Genome.

[39]  R. Shoemaker,et al.  Genome duplication in soybean (Glycine subgenus soja). , 1996, Genetics.

[40]  D. Peterson,et al.  DNA content of heterochromatin and euchromatin in tomato (Lycopersicon esculentum) pachytene chromosomes. , 1996, Genome.

[41]  S. Stack,et al.  Two-dimensional spreads of synaptonemal complexes from solanaceous plants. VI. High-resolution recombination nodule map for tomato (Lycopersicon esculentum). , 1995, Genetics.

[42]  S. Stack,et al.  Two-dimensional spreads of synaptonemal complexes from solanaceous plants. V. Tomato (Lycopersicon esculentum) karyotype and idiogram , 1992 .

[43]  S. Tanksley,et al.  Somatic chromosome karyotype of tomato based on in situ hybridization of the TGRI satellite repeat , 1989 .

[44]  S. Stack.  Heterochromatin, the synaptonemal complex and crossing over. , 1984, Journal of cell science.

[45]  J. S. Heslop-Harrison,et al.  Nuclear dna amounts in angiosperms. , 1976, Philosophical transactions of the Royal Society of London. Series B, Biological sciences.