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Journal of Biochemistry Advance Access originally published online on October 27, 2007
Journal of Biochemistry 2008 143(1):97-105; doi:10.1093/jb/mvm197
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© 2007 The Japanese Biochemical Society.

Reshuffling of the Bacillus subtilis 168 Genome by Multifold Inversion

Azusa Kuroki1,2, Tsutomu Toda3, Kuniko Matsui3, Rie Uotsu-Tomita3, Masaru Tomita2 and Mitsuhiro Itaya2,*

1Graduate School of Media and Governance, Keio University, 5322 Endo, Fujisawa-shi, Kanagawa, 252-8520; 2Institute for Advanced Biosciences, Keio University, 403-1 Nipponkoku, Daihouji, Tsuruoka-shi, Yamagata 997-0017; and 3Mitsubishi Kagaku Institute of Life Sciences, 11 Minamiooya, Machida-shi, Tokyo 194-8511, Japan

*To whom correspondence should be addressed. Tel: +81 (235) 29 0526, Fax: +81 (235) 29 0530, E-mail: mita2001{at}sfc.keio.ac.jp

Received September 6, 2007; Accepted October 8, 2007


  Abstract

The genome of Bacillus subtilis 168 was modified to yield a genome vector for the cloning of DNA several Mb in size. Unlike contemporary plasmid-based vectors, this 4.2 Mb genome vector requires specific in vivo handling protocols because of its large size. Inversion mutagenesis, a method to modify local genome structure without gain or loss of genes, was applied intensively to the B. subtilis genome; this technique made possible both exchange and translocation of designated regions of the genome. This method not only reshuffles the genome of B. subtilis, but can provide insight into the biologic principles underlying genome plasticity.

Key Words: antibiotic resistance, competence, inversion, homologous recombination, translocation, transformation

Abbreviations: Ap, ampicillin; BGM vector, Bacillus subtilis genome vector; BS, Blasticidin S; bsr, Blasticidin S resistance gene; CHEF, contour-clamped homogeneous electric field; Em, erythromycin; erm, erythromycin resistance gene; Nm, neomycin; neo, neomycin resistance gene; Phl, phleomycin; phl, phleomycin resistance gene; Spec, spectinomycin; spc, spectinomycin resistance gene; Tc, tetracycline; tet, tetracycline resistance gene


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