Directed genomic integration in Actinobacillus actinomycetemcomitans: Generation of defined leukotoxin-negative mutants

David J Kolodrubetz, L. H. Phillips, P. J. Ezzo, Ellen B Kraig

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Original languageEnglish
Pages (from-to)2780-2784
Number of pages5
JournalInfection and Immunity
Volume63
Issue number7
StatePublished - 1995

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Aggregatibacter actinomycetemcomitans
Spectinomycin
Plasmids
Genes
Virulence
leukotoxin

ASJC Scopus subject areas

  • Immunology

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Directed genomic integration in Actinobacillus actinomycetemcomitans : Generation of defined leukotoxin-negative mutants. / Kolodrubetz, David J; Phillips, L. H.; Ezzo, P. J.; Kraig, Ellen B.

In: Infection and Immunity, Vol. 63, No. 7, 1995, p. 2780-2784.

Research output: Contribution to journalArticle

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abstract = "To develop targeted gene integration in the periodontal pathogen Actinobacillus actinomycetemcomitans, a ColE1-based, spectinomycin-resistant plasmid containing a segment of the leukotoxin gene was electroporated into strain JP2. In all of the stable spectinomycin-resistant transformants that arose, the plasmid had recombined into the genomic leukotoxin locus since ColE1-based vectors cannot replicate extrachromosomally in A. actinomycetemcomitans. Directed genomic integration was then used to construct a leukotoxin-negative strain by transforming the leukotoxin- producing strain JP2 with a CnlE1-based plasmid containing an internal fragment of the leukotoxin gene. Cytotoxicity assays proved that these transformants had <0.1{\%} of the leukotoxin activity of the parental strain. These results demonstrate that integration-based approaches can be used for generating isogenic mutants in specific virulence genes in A. actinomycetemcomitans.",
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AU - Kraig, Ellen B

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