title: Single Crossover-Mediated Markerless Genome Engineering in Clostridium acetobutylicum.
abstract: A novel genome-engineering tool in Clostridium acetobutylicum was developed based on singlecrossover homologous recombination. A small-sized non-replicable plasmid, pHKO1, was designed for efficient integration into the C. acetobutylicum genome. The integrated pHKO1 plasmid backbone, which included an antibiotic resistance gene, can be excised in vivo by Flp recombinase, leaving a single flippase recognition target sequence in the middle of the targeted gene. Since the pSHL-FLP plasmid, the carrier of the Flp recombinase gene, employed the segregationally unstable pAMβ1 replicon, the plasmid was rapidly cured from the mutant C. acetobutylicum. Consequently, our method makes it easier to engineer C. acetobutylicum.
authors.affiliation: Department of Biotechnology, Graduate School, Korea University, Seoul 02841, Republic of Korea.|R&D Center, GS Caltex Corporation, Daejeon 34122, Republic of Korea.
authors.forename: Sang-Hyun
authors.initials: SH
authors.lastname: Lee
authors.affiliation: Biosystems and Bioengineering Program, University of Science and Technology and Microbiomics and Immunity Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 34141, Republic of Korea.
authors.forename: Hyun Ju
authors.initials: HJ
authors.lastname: Kim
authors.affiliation: R&D Center, GS Caltex Corporation, Daejeon 34122, Republic of Korea.
authors.forename: Yong-An
authors.initials: YA
authors.lastname: Shin
authors.affiliation: Department of Biotechnology, Graduate School, Korea University, Seoul 02841, Republic of Korea.
authors.forename: Kyoung Heon
authors.initials: KH
authors.lastname: Kim
authors.affiliation: Biosystems and Bioengineering Program, University of Science and Technology and Microbiomics and Immunity Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 34141, Republic of Korea.
authors.forename: Sang Jun
authors.initials: SJ
authors.lastname: Lee
mesh_terms: D046969:Clostridium acetobutylicum / Q000235:genetics*; D004254:DNA Nucleotidyltransferases / Q000235:genetics; D020868:Gene Silencing; D005818:Genetic Engineering / Q000379:methods*; D016680:Genome, Bacterial; D059765:Homologous Recombination; D009154:Mutation; D010957:Plasmids
chemical_list: D004254:DNA Nucleotidyltransferases; C045119:FLP recombinase
keywords: Clostridium acetobutylicum; gene inactivation; markerless; single crossover