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.
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Marker excision can be achieved using FLP recombinase or the Cre-lox system.
“This can either be achieved using above mentioned FLP recombinase (Lee et al., 2016) or using the Cre-lox system (Sauer, 1987).”
From Gas Fermentation—A Flexible Platform for Commercial Scale Production of Low-Carbon-Fuels and Chemicals from Waste and Renewable Feedstocks (2016) · cites this paper as Lee et al., 2016
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Self-citation
- Authors
- Lee, S.-H., Kim, H. J., Shin, Y.-A., Kim, K. H., & Lee, S. J.
- Affiliations
- Korea University (KR)
- Korea Research Institute of Bioscience and Biotechnology (KR)
- GS Caltex (South Korea) (KR)
- Corresponding author
- Kim, Kyoung Heon
- Venue
- Journal of Microbiology and Biotechnology
- Published
- 2016-04-28
- Volume
- 26
- Issue
- 4
- Pages
- 725-729
- ISSN
- 1017-7825 (ISSN-L)
- 1738-8872
- OpenAlex
- W2393105221
- Cited by
- 11
- References
- 0
- Citations by year
- 2024: 1 · 2023: 1 · 2021: 3 · 2018: 2 · 2017: 2 · 2016: 2
- FWCI
- 1.04
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Journal of Microbiology and Biotechnology (publishedVersion)
- PubMed (publishedVersion)
- KRIBB Repository (submittedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Microbial Metabolic Engineering and Bioproduction
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Keywords
- Clostridium acetobutylicum, Flp recombinase, plasmid excision, plasmid curing, antibiotic resistance genes, gene integration
- MeSH terms
- DNA Nucleotidyltransferases, Genetic Engineering, Mutation, Plasmids, Genome, Bacterial, Gene Silencing, Clostridium acetobutylicum, Homologous Recombination
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