A mariner transposon vector adapted for mutagenesis in oral streptococci

MicrobiologyOpen

Martin Nilsson¹  Natalia Christiansen¹  Niels Høiby¹  Svante Twetman²  Michael Givskov¹ 

[1] Costerton Biofilm Center, Department of International Health, Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark;Department of Odontology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark

关键词: Biofilm;    extracellular matrix;    glycosyltransferases;    mariner;    mutagenesis;    Streptococcus mutans;    transposon.;   

DOI  :  10.1002/mbo3.171

来源: Wiley

【 摘 要 】

Abstract

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.

【 授权许可】

CC BY   
© 2014 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.