Spontaneous and restriction enzyme-induced chromosomal recombination in mammalian cells

Alan R. Godwin, Roni J. Bollag, Donna Marie Christie, R. Michael Liskay

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Abstract

We have derived Chinese hamster ovary (CHO) cell hybrids containing herpes simplex virus thymidine kinase (tk) heteroalleles for the study of spontaneous and restriction enzyme-induced interchromosomal recombination. These lines allowed us to make a direct comparison between spontaneous intrachromosomal and interchromosomal recombination using the same tk heteroalleles at the same genomic insertion site. We find that the frequency of interchromosomal recombination is less by a factor of at least 5000 than that of intrachromosomal recombination. Our results with mammalian cells differ markedly from results with Saccharomyces cerevisiae, with which similar studies typically give only a 10-to 30-fold difference. Next, to inquire into the fate of double-strand breaks at either of the two different Xho I linker insertion mutations, we electroporated PaeR71 enzyme, an isoschizomer of Xho I, into these hybrids. A priori, these breaks can be repaired either by recombination from the homolog or by end-joining. Despite a predicted bias against recovering end-joining products in our system, all cells characterized by enzyme-induced resistance to hypoxanthine/aminopter- in/thymidine were, in fact, due to nonhomologous recombination or end- joining. These results are in agreement with other studies that used extrachromosomal sequences to examine the relative efficiencies of end- joining and homologous recombination in mammalian cells, but are in sharp contrast to results of analogous studies in S. cerevisiae, wherein only products of homologous events are detected.

Original languageEnglish (US)
Pages (from-to)12554-12558
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume91
Issue number26
DOIs
StatePublished - Dec 20 1994

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Keywords

  • double- strand break
  • interchromosomal recombination
  • intrachromosomal recombination

ASJC Scopus subject areas

  • General

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