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Applied and Environmental Microbiology, May 2002, p. 2359-2367, Vol. 68, No. 5
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.5.2359-2367.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Cre-loxP Recombination System for Large Genome Rearrangements in Lactococcus lactis{dagger}

Nathalie Campo,1 Marie-Line Daveran-Mingot,1 Kees Leenhouts ,2,{ddagger} Paul Ritzenthaler,1* and Pascal Le Bourgeois1

Laboratoire de Microbiologie et Génétique Moléculaire du CNRS (UMR5100), Université Paul Sabatier, 31062 Toulouse, France,1 Department of Genetics, Biomolecular Sciences and Biotechnology Institute, University of Groningen, Haren, The Netherlands2

Received 6 December 2001/ Accepted 26 February 2002

We have used a new genetic strategy based on the Cre-loxP recombination system to generate large chromosomal rearrangements in Lactococcus lactis. Two loxP sites were sequentially integrated in inverse order into the chromosome either at random locations by transposition or at fixed points by homologous recombination. The recombination between the two chromosomal loxP sites was highly efficient (approximately 1 x 10-1/cell) when the Cre recombinase was provided in trans, and parental- or inverted-type chromosomal structures were isolated after removal of the Cre recombinase. The usefulness of this approach was demonstrated by creating three large inversions of 500, 1,115, and 1,160 kb in size that modified the lactococcal genome organization to different extents. The Cre-loxP recombination system described can potentially be used for other gram-positive bacteria without further modification.

* Corresponding author. Mailing address: LMGM du CNRS, Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex, France. Phone: (33) 561 33 58 25. Fax: (33) 561 33 58 86. E-mail: ritzenth{at}ibcg.biotoul.fr.

{dagger} This paper is dedicated to the memory of Patrick Duwat (5 January 2000).

{ddagger} Present address: Biomade Technology Foundation, Mijenbergh 4, 9747 AG Groningen, The Netherlands.

Applied and Environmental Microbiology, May 2002, p. 2359-2367, Vol. 68, No. 5
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.5.2359-2367.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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