A method to invert DNA segments of the Bacillus subtilis 168 genome by recombination between two homologous sequences

Tsutomu Toda, Teruo Tanaka, Mitsuhiro Itaya

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

We developed a method that allows rapid isolation of mutant Bacillus subtilis 168 carrying an inversion of a specific DNA segment of the genome. Two incomplete neomycin resistance gene cassettes were integrated at both ends of the 1652-kb segment to be inverted. Reciprocal recombination within the 590-bp homologous region of these two cassettes created an intact neomycin resistance gene with concomitant inversion of the 1652- kb segment flanked by the two cassettes. Structure of the mutant genome was verified by analyzing the physical map for rare cutting endonucleases, SfiI, NotI, I- CeuI, and I-SceI. The inversion rate was estimated to be 6.9 ± 1.4 x 10- 8/cell/cell division at 37°C. The method should be in principle applicable not only to other regions of the B. subtilis genome but also to other bacterial genomes.

Original languageEnglish
Pages (from-to)773-778
Number of pages6
JournalBioscience, Biotechnology and Biochemistry
Volume60
Issue number5
Publication statusPublished - 1996 May
Externally publishedYes

Fingerprint

Bacilli
Sequence Homology
Bacillus subtilis
sequence homology
Genetic Recombination
Neomycin
Genes
Genome
genome
neomycin
DNA
Bacterial Genomes
Cell Division
mutants
physical chromosome mapping
methodology
rapid methods
cell division
genes
Cells

Keywords

  • bacterial genome technology
  • DNA inversion
  • neomycin
  • rare-cutting endonucleases
  • recA

ASJC Scopus subject areas

  • Food Science
  • Applied Microbiology and Biotechnology
  • Chemistry (miscellaneous)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Biotechnology

Cite this

A method to invert DNA segments of the Bacillus subtilis 168 genome by recombination between two homologous sequences. / Toda, Tsutomu; Tanaka, Teruo; Itaya, Mitsuhiro.

In: Bioscience, Biotechnology and Biochemistry, Vol. 60, No. 5, 05.1996, p. 773-778.

Research output: Contribution to journalArticle

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