Transduction-mediated transfer of unmarked deletion and point mutations through use of counterselectable suicide vectors

Young Kang Ho Young Kang, C. M. Dozois, S. A. Tinge, Ho Lee Tae Ho Lee, R. Curtiss

Research output: Contribution to journalArticle

42 Citations (Scopus)

Abstract

A challenge in strain construction is that unmarked deletion and nucleotide substitution alleles generally do not confer selectable phenotypes. We describe here a rapid and efficient strategy for transferring such alleles via generalized transduction. The desired allele is first constructed and introduced into the chromosome by conventional allelic-exchange methods. The suicide vector containing the same allele is then integrated into the mutant chromosome, generating a tandem duplication homozygous for that allele. The resulting strain is used as a donor for transductional crosses, and selection is made for a marker carried by the integrated suicide vector. Segregation of the tandem duplication results in haploid individuals, each of which carries the desired allele. To demonstrate this mutagenesis strategy, we used bacteriophage P22HTint for generalized transduction-mediated introduction of unmarked mutations to Salmonella enterica serovar Typhimurium. This method is applicable to any species for which generalized transduction is established.

Original languageEnglish (US)
Pages (from-to)307-312
Number of pages6
JournalJournal of Bacteriology
Volume184
Issue number1
DOIs
StatePublished - 2002
Externally publishedYes

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Sequence Deletion
Point Mutation
Suicide
Alleles
Chromosomes
Salmonella enterica
Haploidy
Mutagenesis
Bacteriophages
Nucleotides
Phenotype
Mutation

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Immunology

Cite this

Transduction-mediated transfer of unmarked deletion and point mutations through use of counterselectable suicide vectors. / Ho Young Kang, Young Kang; Dozois, C. M.; Tinge, S. A.; Tae Ho Lee, Ho Lee; Curtiss, R.

In: Journal of Bacteriology, Vol. 184, No. 1, 2002, p. 307-312.

Research output: Contribution to journalArticle

Ho Young Kang, Young Kang ; Dozois, C. M. ; Tinge, S. A. ; Tae Ho Lee, Ho Lee ; Curtiss, R. / Transduction-mediated transfer of unmarked deletion and point mutations through use of counterselectable suicide vectors. In: Journal of Bacteriology. 2002 ; Vol. 184, No. 1. pp. 307-312.
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