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• W2096557809 endingPage "7307" @default.
• W2096557809 startingPage "7298" @default.
• W2096557809 abstract "ABSTRACT Transposon mutagenesis in bacteria generally requires efficient delivery of a transposon suicide vector to allow the selection of relatively infrequent transposition events. We have developed an IS 903 -based transposon mutagenesis system for diverse gram-negative bacteria that is not limited by transfer efficiency. The transposon, IS 903φkan , carries a cryptic kan gene, which can be expressed only after successful transposition. This allows the stable introduction of the transposon delivery vector into the host. Generation of insertion mutants is then limited only by the frequency of transposition. IS 903φkan was placed on an IncQ plasmid vector with the transposase gene located outside the transposon and expressed from isopropyl-β- d -thiogalactopyranoside (IPTG)-inducible promoters. After transposase induction, IS 903φkan insertion mutants were readily selected in Escherichia coli by their resistance to kanamycin. We used IS 903φkan to isolate three catalase-deficient mutants of the periodontal pathogen Actinobacillus actinomycetemcomitans from a library of random insertions. The mutants display increased sensitivity to hydrogen peroxide, and all have IS 903φkan insertions within an open reading frame whose predicted product is closely related to other bacterial catalases. Nucleotide sequence analysis of the catalase gene (designated katA ) and flanking intergenic regions also revealed several occurrences of an 11-bp sequence that is closely related to the core DNA uptake signal sequence for natural transformation of Haemophilus influenzae . Our results demonstrate the utility of the IS 903φkan mutagenesis system for the study of A. actinomycetemcomitans . Because IS 903φkan is carried on a mobilizable, broad-host-range IncQ plasmid, this system is potentially useful in a variety of bacterial species." @default.
• W2096557809 created "2016-06-24" @default.
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• W2096557809 date "1999-12-01" @default.
• W2096557809 modified "2023-09-27" @default.
• W2096557809 title "Direct Selection of IS <i>903</i> Transposon Insertions by Use of a Broad-Host-Range Vector: Isolation of Catalase-Deficient Mutants of <i>Actinobacillus actinomycetemcomitans</i>" @default.
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• W2096557809 doi "https://doi.org/10.1128/jb.181.23.7298-7307.1999" @default.
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