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. 2015 Jun 11;14:80. doi: 10.1186/s12934-015-0269-6

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© Tajima et al. 2015

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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Scheme of A. succinogenes PCK expression cassette integration into the poxB locus using the λ Red- and Int/Xis-dependent systems. a Construction of A. succinogenes PCK expression cassette, b introduction of the PCK expression cassette into poxB locus on chromosome by λ Red-dependent recombination system using pRSFRedTER, c replacement of the poxB gene of the ES06 strain with the PCK expression cassette, and d removal of the kanamycin resistance gene from chromosome by the λ Int/Xis-dependent excision system using pRSF-P _ara-IX. These genetic manipulations leave a 31-bp attB _λ site (5′-TGAAGCCTGC TTTTTTATAC TAACTTGAGC G-3′) in the chromosome.