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GENE CLUSTER OR OPERON DESIGN BY ORDERED GENE ASSEMBLY IN BACILLUS SUBTILIS (OGAB) METHOD, pp. 153-168 $0.00
Authors:  Kenji Tsuge, Mitsuhiro Itaya, Institute for Advanced Biosciences, Keio University, Yamagata, Japan, and others
Abstract:
In the post genome sequence era, metabolic engineering conducted by a number of
genes is attracting great attention for not only basic research but also for application in
production of beneficial materials for our lives. To carry this out, methods to handle a
number of genes at a time are a prerequisite. However, the poor ability of Escherichia
coli-based genetic methods to manipulate the reconstruction makes the steps timeconsuming
and laborious. Bacillus subtilis, a gram-positive soil bacterium, develops
natural competence in certain physiological conditions. In the competence, this bacterium
can take up exogenous DNAs, and can recombine these DNAs with the genome if there is
a homologous sequence. By using this trait, a novel method to carry out gene assembly
quickly and with greatest fidelity using B. subtilis was developed. The method, named
Ordered Gene Assembly in B. subtilis (OGAB), facilitates the connection of more than
10 DNA fragments in plasmid in one-step using an intrinsic B. subtilis plasmid
transformation system. In this chapter, the principle of and actual protocol for the OGAB
method are written. Moreover, two types of gene assembly in terms of metabolic
engineering are also described. 


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GENE CLUSTER OR OPERON DESIGN BY ORDERED GENE ASSEMBLY IN BACILLUS SUBTILIS (OGAB) METHOD, pp. 153-168