Single crossover-mediated markerless genome engineering in clostridium acetobutylicum

Sang Hyun Lee, Hyun Ju Kim, Yong An Shin, Kyoung Heon Kim, Sang Jun Lee

Research output: Contribution to journalArticlepeer-review

9 Citations (Scopus)


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.

Original languageEnglish
Pages (from-to)725-729
Number of pages5
JournalJournal of Microbiology and Biotechnology
Issue number4
Publication statusPublished - 2016 Apr 28


  • Clostridium acetobutylicum
  • Gene inactivation
  • Markerless
  • Single crossover

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology


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