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Applied and Environmental Microbiology, May 2009, p. 2638-2642, Vol. 75, No. 9
0099-2240/09/$08.00+0     doi:10.1128/AEM.02214-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Generation of Single-Copy Transposon Insertions in Clostridium perfringens by Electroporation of Phage Mu DNA Transposition Complexes{triangledown}

A. Lanckriet,1* L. Timbermont,1 L. J. Happonen,2 M. I. Pajunen,2,3 F. Pasmans,1 F. Haesebrouck,1 R. Ducatelle,1 H. Savilahti,2,3 and F. Van Immerseel1

Department of Pathology, Bacteriology and Avian Diseases, Research Group for Veterinary Public Health and Zoonoses, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, B-9820 Merelbeke, Belgium,1 Research Program in Cellular Biotechnology, Institute of Biotechnology, Viikki Biocenter, P.O. Box 56, Viikinkaari 9, FIN-00014 University of Helsinki, Finland,2 Division of Genetics and Physiology, Department of Biology, Vesilinnantie 5, FIN-20014 University of Turku, Finland3

Received 25 September 2008/ Accepted 22 February 2009

Transposon mutagenesis is a tool that is widely used for the identification of genes involved in the virulence of bacteria. Until now, transposon mutagenesis in Clostridium perfringens has been restricted to the use of Tn916-based methods with laboratory reference strains. This system yields primarily multiple transposon insertions in a single genome, thus compromising its use for the identification of virulence genes. The current study describes a new protocol for transposon mutagenesis in C. perfringens, which is based on the bacteriophage Mu transposition system. The protocol was successfully used to generate a single-insertion mutant library both for a laboratory strain and for a field isolate. Thus, it can be used as a tool in large-scale screening to identify virulence genes of C. perfringens.

* Corresponding author. Mailing address: Department of Pathology, Bacteriology and Avian Diseases, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, B-9820 Merelbeke, Belgium. Phone: (32) 09 264 74 48. Fax: (32) 09 264 74 94. E-mail: anouk.lanckriet{at}UGent.be

{triangledown} Published ahead of print on 6 March 2009.

Applied and Environmental Microbiology, May 2009, p. 2638-2642, Vol. 75, No. 9
0099-2240/09/$08.00+0     doi:10.1128/AEM.02214-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



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