Versatile nourseothricin and streptomycin/spectinomycin resistance gene cassettes and their use in chromosome integration vectors

Stephanie S. Lehman, Katherine M. Mladinich, Angkana Boonyakanog, Takehiko Mima, Rox Ann R Karkhoff-Schweizer, Herbert P. Schweizer

Research output: Contribution to journalArticle

Abstract

An obstacle for the development of genetic systems for many bacteria is the limited number of antibiotic selection markers, especially for bacteria that are intrinsically antibiotic resistant or where utilization of such markers is strictly regulated. Here we describe the development of versatile cassettes containing nourseothricin, streptomycin/spectinomycin, and spectinomycin selection markers. The antibiotic resistance genes contained on these cassettes are flanked by loxP sites with allow their in vivo excision from the chromosome of target bacteria using Cre recombinase. The respective selection marker cassettes were used to derive mini-Tn7 elements that can be used for single-copy insertion of genes into bacterial chromosomes. The utility of the selection markers was tested by insertion of the resulting mini-Tn7 elements into the genomes of Burkholderia thailandensis and B. pseudomallei efflux pump mutants susceptible to aminoglycosides, aminocyclitols, and streptothricins, followed by Cre-mediated antibiotic resistance marker excision. The versatile nourseothricin, streptomycin/spectinomycin and spectinomycin resistance loxP cassette vectors described here extend the repertoire of antibiotic selection markers for genetic manipulation of diverse bacteria that are susceptible to aminoglycosides and aminocyclitols.

Original languageEnglish
Pages (from-to)8-13
Number of pages6
JournalJournal of Microbiological Methods
Volume129
DOIs
Publication statusPublished - Oct 1 2016
Externally publishedYes

Fingerprint

Streptothricins
Spectinomycin
Streptomycin
Chromosomes
Bacteria
Aminoglycosides
Microbial Drug Resistance
Anti-Bacterial Agents
Genes
Bacterial Chromosomes
Burkholderia
Insertional Mutagenesis
Genetic Markers
Genome

Keywords

  • Antibiotics
  • Cassette
  • Mini-Tn7
  • Nourseothricin
  • Selection
  • Spectinomycn
  • Streptomycin

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology
  • Microbiology (medical)

Cite this

Versatile nourseothricin and streptomycin/spectinomycin resistance gene cassettes and their use in chromosome integration vectors. / Lehman, Stephanie S.; Mladinich, Katherine M.; Boonyakanog, Angkana; Mima, Takehiko; Karkhoff-Schweizer, Rox Ann R; Schweizer, Herbert P.

In: Journal of Microbiological Methods, Vol. 129, 01.10.2016, p. 8-13.

Research output: Contribution to journalArticle

Lehman, Stephanie S. ; Mladinich, Katherine M. ; Boonyakanog, Angkana ; Mima, Takehiko ; Karkhoff-Schweizer, Rox Ann R ; Schweizer, Herbert P. / Versatile nourseothricin and streptomycin/spectinomycin resistance gene cassettes and their use in chromosome integration vectors. In: Journal of Microbiological Methods. 2016 ; Vol. 129. pp. 8-13.
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