A method to invert DNA segments of the Bacillus subtilis 168 genome by recombination between two homologous sequences.
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 degrees C. The method should be in principle applicable not only to other regions of the B, subtilis genome but also to other bacterial genomes.
Toda
T
T
Mitsubishi Kasei Institute of Life Sciences, Tokyo, Japan.
Tanaka
T
T
Itaya
M
M
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001412:Bacillus subtilis / Q000187:drug effects / Q000235:genetics*; D015139:Blotting, Southern; D007446:Chromosome Inversion; D002876:Chromosomes, Bacterial / Q000235:genetics; D004262:DNA Restriction Enzymes / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics*; D004352:Drug Resistance, Microbial / Q000235:genetics; D016680:Genome, Bacterial; D016254:Mutagenesis, Insertional; D009154:Mutation / Q000235:genetics; D009355:Neomycin / Q000494:pharmacology; D011995:Recombination, Genetic / Q000187:drug effects; D012689:Sequence Homology, Nucleic Acid
D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D004262:DNA Restriction Enzymes; D009355:Neomycin