Abstract
Genome rearrangement is an increasingly important technique to facilitate the understanding of genome functions. A Cre/loxP-mediated deletion system for large-scale genome rearrangements in Corynebacterium glutamicum was developed. By comparative analysis of C. glutamicum R and C. glutamicum 13032 genomes, distinct 14.5-kb and 56-kb regions not essential for cell survival were identified and targeted for deletion. By homologous recombination, loxP sites were integrated at each end of the target region. Deletions between the two chromosomal loxP sites in the presence of Cre recombinase were highly efficient. Accurate deletion was observed in all 96 Cre-expressing strains tested. These deletions represent the largest genomic excisions in C. glutamicum reported to date. Despite the loss of 11 and 58 predicted ORF(s), respectively, upon the deletion of the14.5-kb and 56-kb regions, the cells still exhibited normal growth under standard laboratory conditions. Based on the precision of its deletion, the Cre/loxP system provides a new, efficient genome rearrangement technique for studying C. glutamicum.
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Acknowledgments
We wish to thank Dr Ken-ichi Yamamura for plasmid pBS185. This study was carried out as a part of The Project for Development of a Technological Infrastructure for Industrial Bioprocesses on R&D of New Industrial Science and Technology Frontiers by Ministry of Economy, Trade & Industry (METI), and entrusted by New Energy and Industrial Technology Development Organization (NEDO).
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Suzuki, N., Tsuge, Y., Inui, M. et al. Cre/loxP-mediated deletion system for large genome rearrangements in Corynebacterium glutamicum. Appl Microbiol Biotechnol 67, 225–233 (2005). https://doi.org/10.1007/s00253-004-1772-6
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DOI: https://doi.org/10.1007/s00253-004-1772-6