• Journal of Life Science
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• v.16 no.7 s.80
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• pp.1112-1118
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• 2006

Bordetella bronchiseptica, the agent of swine atrophic rhinitis and kennel cough in dogs, is a mucosal pathogen and produces the hydroxamate type alcaligin siderophore under iron-limited conditions. Genes involved in alcaligin siderophore biosynthesis are contained in an alcABCDE operon. In order to provide direct evidence for the role of AlcA in alcaligin biosynthesis, we needed a B. bronchiseptica mutant carrying alcA gene deletion. A 0.6 kb alcA 5'-flanking and 0.7kb 3'-flanking DNA fragments were PCR amplified with the use of pCP1.11 as a template DNA. The 5'-and 3'-flanking DNA fragments were joined in a suicide plasmid, resulting in a recombinant suicide plasmid pDM1. After introduction of pDM1 into B. bronchiseptica by conjugation, the allelic exchange technique was performed and a B. bronchiseptica alcA deletion mutant, named B. bronchiseptica H1, was obtained. The mutant strain produced reduced amount of siderophore as expected. When a plasmid containing complete alcA gene was transformed back into the mutant, the complemented mutant recovered ability of siderophore production. These results indicated that AlcA is one of essential components for the alcaligin siderophore biosynthesis. The mutant strains obtained in this study will be used in the further studies for the biochemical function of AlcA.