Intrachromosomal recombination between well-separated, homologous sequences in mammalian cells.
In the present study, we investigated intrachromosomal homologous recombination in a murine hybridoma in which the recipient for recombination, the haploid, endogenous chromosomal immunoglobulin mu-gene bearing a mutation in the constant (Cmu) region, was separated from the integrated single copy wild-type donor Cmu region by approximately 1 Mb along the hybridoma chromosome. Homologous recombination between the donor and recipient Cmu region occurred with high frequency, correcting the mutant chromosomal mu-gene in the hybridoma. This enabled recombinant hybridomas to synthesize normal IgM and to be detected as plaque-forming cells (PFC). Characterization of the recombinants revealed that they could be placed into three distinct classes. The generation of the class I recombinants was consistent with a simple unequal sister chromatid exchange (USCE) between the donor and recipient Cmu region, as they contained the three Cmu-bearing fragments expected from this recombination, the original donor Cmu region along with both products of the single reciprocal crossover. However, a simple mechanism of homologous recombination was not sufficient in explaining the more complex Cmu region structures characterizing the class II and class III recombinants. To explain these recombinants, a model is proposed in which unequal pairing between the donor and recipient Cmu regions located on sister chromatids resulted in two crossover events. One crossover resulted in the deletion of sequences from one chromatid forming a DNA circle, which then integrated into the sister chromatid by a second reciprocal crossover.
Baker
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MD
Department of Pathobiology, University of Guelph, Guelph, Ontario N1G 2W1, Canada. mbaker@ovcnet.uoguelph.ca
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Beatty
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D000818:Animals; D002460:Cell Line; D002875:Chromosomes / Q000235:genetics; D004247:DNA / Q000235:genetics*; D006825:Hybridomas; D007148:Immunoglobulin mu-Chains / Q000235:genetics; D017404:In Situ Hybridization, Fluorescence; D008322:Mammals; D009154:Mutation; D011995:Recombination, Genetic
D007148:Immunoglobulin mu-Chains; D004247:DNA