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Plant Molecular Biology

, Volume 32, Issue 5, pp 901–913 | Cite as

Single-site manipulation of tomato chromosomes in vitro and in vivo using Cre-lox site-specific recombination

  • Jeroen Stuurman
  • Marianne J. de Vroomen
  • H. John J. Nijkamp
  • Mark J. J. van Haaren
Research Article

Abstract

With the aim of developing new techniques for physical and functional genome analysis, we have introduced the Cre-lox site-specific recombination system into the cultivated tomato (Lycopersicon esculentum). Local transposition of a Ds(lox) transposable element from a T-DNA(lox) on the long arm of chromosome 6 was used to position pairs of lox sites on different closely linked loci. In vitro Cre-lox recombination between chromosomal lox sites and synthetic lox oligonucleotides cleaved the 750 Mb tomato genome with 34 pb specificity to release unique 65 kb and 130 kb fragments of chromosome 6. Parallel in vitro experiments on Saccharomyces cerevisiae chromosomes show the efficiency of cleavage to be 50% per chromosomal lox site at maximum. By expressing the Cre recombinase in tomato under control of a constitutive CaMV 35S promoter, efficient and specific somatic and germinal in planta inversion of the 130 kb fragment is demonstrated. The combined use of in vitro and in vivo recombination on genetically mapped lox sites will provide new possibilities for long range restriction mapping and in vivo manipulation of selected tomato genome segments.

Key words

Ac/Ds chromosomal rearrangements Cre-lox meganucleases tomato 

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Copyright information

© Kluwer Academic Publishers 1996

Authors and Affiliations

  • Jeroen Stuurman
    • 1
  • Marianne J. de Vroomen
    • 1
  • H. John J. Nijkamp
    • 1
  • Mark J. J. van Haaren
    • 1
  1. 1.Institute for Molecular Biological Sciences, Department of GeneticsFree University AmsterdamAmsterdamNetherlands

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