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Region Hovedstaden - en del af Københavns Universitetshospital
Udgivet

A mariner transposon vector adapted for mutagenesis in oral streptococci

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DOI

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  • Martin Nilsson
  • Natalia Christiansen
  • Niels Høiby
  • Svante Twetman
  • Michael Givskov
  • Tim Tolker-Nielsen
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This article describes the construction and characterization of a mariner-based transposon vector designed for use in oral streptococci, but with a potential use in other Gram-positive bacteria. The new transposon vector, termed pMN100, contains the temperature-sensitive origin of replication repATs-pWV01, a selectable kanamycin resistance gene, a Himar1 transposase gene regulated by a xylose-inducible promoter, and an erythromycin resistance gene flanked by himar inverted repeats. The pMN100 plasmid was transformed into Streptococcus mutans UA159 and transposon mutagenesis was performed via a protocol established to perform high numbers of separate transpositions despite a low frequency of transposition. The distribution of transposon inserts in 30 randomly picked mutants suggested that mariner transposon mutagenesis is unbiased in S. mutans. A generated transposon mutant library containing 5000 mutants was used in a screen to identify genes involved in the production of sucrose-dependent extracellular matrix components. Mutants with transposon inserts in genes encoding glycosyltransferases and the competence-related secretory locus were predominantly found in this screen.

OriginalsprogEngelsk
TidsskriftMicrobiologyOpen
Vol/bind3
Udgave nummer3
Sider (fra-til)333-40
Antal sider8
DOI
StatusUdgivet - jun. 2014

ID: 44828683