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Research article

J. Microbiol. Biotechnol. 2015; 25(10): 1702-1708

Published online October 28, 2015 https://doi.org/10.4014/jmb.1504.04070

Copyright © The Korean Society for Microbiology and Biotechnology.

A New Shuttle Plasmid That Stably Replicates in Clostridium acetobutylicum

Sang-Hyun Lee 1, 2, Min-A Kwon 2, Sunwha Choi 2, Sooah Kim 1, Jungyeon Kim 1, Yong-An Shin 2 and Kyoung Heon Kim 1*

1Department of Biotechnology, Graduate School, Korea University, Seoul 136-713, Republic of Korea, 2R&D Center, GS Caltex Corporation, Daejeon 305-380, Republic of Korea

Received: April 27, 2015; Accepted: May 28, 2015

Abstract

We have developed a new shuttle plasmid, designated as pLK1-MCS that can replicate in both
Clostridium acetobutylicum and Escherichia coli, by combining the pUB110 and pUC19 plasmids.
Plasmid pLK1-MCS replicated more stably than previously reported plasmids containing
either the pIM13 or the pAMβ1 replicon in the absence of antibiotic selective pressure. The
transfer frequency of pLK1-MCS into C. acetobutylicum was similar to the transfer frequency of
other shuttle plasmids. We complemented C. acetobutylicum ML1 (that does not produce
solvents such as acetone, butanol, and ethanol owing to loss of the megaplasmid pSOL1
harboring the adhE1-ctfAB-adc operon) by introducing pLK1-MCS carrying the adhE1-ctfAB-adc
operon into C. acetobutylicum ML1. The transformed cells were able to resume anaerobic
solvent production, indicating that the new shuttle plasmid has the potential for practical use
in microbial biotechnology.

Keywords: Clostridium acetobutylicum, shuttle plasmid, segregational stability