Theta-type DNA replication stimulates homologous recombination in the Bacillus subtilis chromosome

Authors

  • Françoise Morel-Deville,

    1. Laboratoire de Génétique Microbienne, Institut National de la Recherche Agronomique, Domaine de Vilvert, 78352 Jouy en Josas Cedex, France
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  • S. Dusko Ehrlich

    1. Laboratoire de Génétique Microbienne, Institut National de la Recherche Agronomique, Domaine de Vilvert, 78352 Jouy en Josas Cedex, France
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Françoise Morel-Deville Laboratoire de Recherche sur la Viande, INRA, Domaine de Vilvert, 78352 Jouy-en-Josas Cedex, France. E-mail fmorel@biotec.jouy.inra.fr; Tel. (1) 34 65 22 92; Fax (1) 34 65 21 05

Abstract

To test the effects of theta-type replication on homologous DNA recombination, we integrated in the chromosome of Bacillus subtilis a structure comprising a conditional replication region and direct repeats of ∼ 4 kb. The replicon was derived from a broad-host-range plasmid, pAMβ1, which replicates by a unidirectional theta mechanism and is thermosensitive. The direct repeats were derived from plasmid pBR322 and flanked the chloramphenicol-resistance gene of plasmid pC194. Recombination between the repeats could therefore lead to a loss of the resistance gene or the appearance of additional repeats. The integrated replicon was active at the permissive temperature, and ∼ 25% of the integrated plasmids could be isolated as Y-shaped molecules after restriction, having a branch at the replication origin. Replicon activity stimulated recombination four- to fivefold, as estimated from the proportion of chloramphenicol-sensitive cells at the restrictive and permissive temperature, and also led to the appearance of additional direct repeats. We conclude that theta-type replication stimulates homologous recombination and suggest that many or even most recombination events between long homologous sequences present in a bacterial genome may be the consequence of DNA replication.

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