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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

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

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Tidsskriftspublikasjon
Sammendrag
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.
Språk
Engelsk
Forfatter(e)
Institusjon(er)
  • Universität Bielefeld
  • SINTEF Industri / Bioteknologi og nanomedisin
  • Norges teknisk-naturvitenskapelige universitet
År
2016
Publisert i
Frontiers in Microbiology
ISSN
1664-302X
Forlag
Frontiers Media S.A.
Årgang
7