Genome-based genetic tool development for Bacillus methanolicus: theta- and rolling circle-replicating plasmids for inducible gene expression and application to methanol-based cadaverine production

Irla M, Heggeset TM, Naerdal I, Paul L, Haugen T, Le SB, Brautaset T, Wendisch VF (2016)
Frontiers in Microbiology 7: 1481.

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Abstract
Bacillus methanolicus is a thermophilic methylotroph able to overproduce amino acids from methanol, a substrate not used for human or animal nutrition. Based on our previous RNA-seq analysis a mannitol inducible promoter and a putative mannitol activator gene mtlR were identified. The mannitol inducible promoter was applied for controlled gene expression using fluorescent reporter proteins and a flow cytometry analysis, and improved by changing the -35 promoter region and by co-expression of the mtlR regulator gene. For independent complementary gene expression control, the heterologous xylose-inducible system from B. megaterium was employed and a two-plasmid gene expression system was developed. Four different replicons for expression vectors were compared with respect to their copy number and stability. As an application example, methanol-based production of cadaverine was shown to be improved from 11.3 to 17.5 g/L when a heterologous lysine decarboxylase gene cadA was expressed from a theta-replicating rather than a rolling-circle replicating vector. The current work on inducible promoter systems and compatible theta- or rolling circle-replicating vectors is an important extension of the poorly developed B. methanolicus genetic toolbox, valuable for genetic engineering and further exploration of this bacterium.
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Article Processing Charge funded by the Deutsche Forschungsgemeinschaft and the Open Access Publication Fund of Bielefeld University.
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Irla M, Heggeset TM, Naerdal I, et al. Genome-based genetic tool development for Bacillus methanolicus: theta- and rolling circle-replicating plasmids for inducible gene expression and application to methanol-based cadaverine production. Frontiers in Microbiology. 2016;7: 1481.
Irla, M., Heggeset, T. M., Naerdal, I., Paul, L., Haugen, T., Le, S. B., Brautaset, T., et al. (2016). Genome-based genetic tool development for Bacillus methanolicus: theta- and rolling circle-replicating plasmids for inducible gene expression and application to methanol-based cadaverine production. Frontiers in Microbiology, 7: 1481.
Irla, M., Heggeset, T. M., Naerdal, I., Paul, L., Haugen, T., Le, S. B., Brautaset, T., and Wendisch, V. F. (2016). Genome-based genetic tool development for Bacillus methanolicus: theta- and rolling circle-replicating plasmids for inducible gene expression and application to methanol-based cadaverine production. Frontiers in Microbiology 7:1481.
Irla, M., et al., 2016. Genome-based genetic tool development for Bacillus methanolicus: theta- and rolling circle-replicating plasmids for inducible gene expression and application to methanol-based cadaverine production. Frontiers in Microbiology, 7: 1481.
M. Irla, et al., “Genome-based genetic tool development for Bacillus methanolicus: theta- and rolling circle-replicating plasmids for inducible gene expression and application to methanol-based cadaverine production”, Frontiers in Microbiology, vol. 7, 2016, : 1481.
Irla, M., Heggeset, T.M., Naerdal, I., Paul, L., Haugen, T., Le, S.B., Brautaset, T., Wendisch, V.F.: Genome-based genetic tool development for Bacillus methanolicus: theta- and rolling circle-replicating plasmids for inducible gene expression and application to methanol-based cadaverine production. Frontiers in Microbiology. 7, : 1481 (2016).
Irla, Marta, Heggeset, Tonje M., Naerdal, Ingemar, Paul, Lidia, Haugen, Tone, Le, Simone B., Brautaset, Trygve, and Wendisch, Volker F. “Genome-based genetic tool development for Bacillus methanolicus: theta- and rolling circle-replicating plasmids for inducible gene expression and application to methanol-based cadaverine production”. Frontiers in Microbiology 7 (2016): 1481.
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