Pulsed field gel electrophoresis and physical mapping of large DNA fragments in the Tm-2a region of chromosome 9 in tomato

Martin W. Ganal, Nevin D. Young, Steven D. Tanksley

Research output: Contribution to journalArticle

116 Scopus citations

Abstract

A method has been developed which allows the isolation of very high molecular weight DNA (>2 million bp) from leaf protoplasts of tomato (Lycopersicon esculentum). The DNA isolated in this manner was digested in agarose with rare-cutting restriction enzymes and separated by pulsed field gel electrophoresis. The size range of the reslting fragments was determined by hybridization to a number of single copy clones and the suitability of these enzymes for the mapping of large DNA fragments was evaluated. Furthermore, five genetically tightly linked single copy clones have been used to begin the construction of a physical map in a region of the genome containing the Tm-2a gene which confers resistance to tobacco mosaic virus. Two of the five clones were found to be on the same 560 kb SalI fragment and therefore are no further apart than that distance. The remaining three markers are distributed over at least 3 million bp, so that the total minimum physical distance of that cluster is at least 4 million bp. The results are discussed with respect to correlations between recombination frequencies and physical distance as well as physical mapping large regions of a complex plant genome like tomato.

Original languageEnglish (US)
Pages (from-to)395-400
Number of pages6
JournalMGG Molecular & General Genetics
Volume215
Issue number3
DOIs
StatePublished - Feb 1 1989

Keywords

  • Lycopersicon esculentum
  • Physical mapping
  • Recombination
  • Restriction fragment length polymorphism
  • Tobacco mosaic virus

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