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Mol Cell Biol. 1995 Apr;15(4):1968-73.

Induction of homologous recombination in mammalian chromosomes by using the I-SceI system of Saccharomyces cerevisiae.

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  • 1Unité de Biologie moléculaire du Développement, Institut Pasteur, Université Pierre et Marie Curie, Paris, France.

Abstract

The mitochondrial intron-encoded endonuclease I-SceI of Saccharomyces cerevisiae has an 18-bp recognition sequence and, therefore, has a very low probability of cutting DNA, even within large genomes. We demonstrate that double-strand breaks can be initiated by the I-SceI endonuclease at a predetermined location in the mouse genome and that the breaks can be repaired with a donor molecule homologous regions flanking the breaks. This induced homologous recombination is approximately 2 orders of magnitude more frequent than spontaneous homologous recombination and at least 10 times more frequent than random integration near an active promoter. As a consequence of induced homologous recombination, a heterologous novel sequence can be inserted at the site of the break. This recombination can occur at a variety of chromosomal targets in differentiated and multipotential cells. These results demonstrate homologous recombination involving chromosomal DNA by the double-strand break repair mechanism in mammals and show the usefulness of very rare cutter endonucleases, such as I-SceI, for designing genome rearrangements.

PMID:
7891691
[PubMed - indexed for MEDLINE]
PMCID:
PMC230423
Free PMC Article
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