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@ARTICLE{Kortmann:173351,
      author       = {Kortmann, Maike and Kuhl, Vanessa and Klaffl, Simon and
                      Bott, Michael},
      title        = {{A} chromosomally encoded {T}7 {RNA} polymerase-dependent
                      gene expression system for {C}orynebacterium glutamicum:
                      construction and comparative evaluation at the single-cell
                      level},
      journal      = {Microbial biotechnology},
      volume       = {8},
      number       = {2},
      issn         = {1751-7915},
      address      = {Oxford},
      publisher    = {Wiley-Blackwell},
      reportid     = {FZJ-2014-06761},
      pages        = {253–265},
      year         = {2015},
      note         = {Biotechnologie 1},
      abstract     = {Corynebacterium glutamicum has become a favourite model
                      organism in white biotechnology. Nevertheless, only few
                      systems for the regulatable (over)expression of homologous
                      and heterologous genes are currently available, all of which
                      are based on the endogenous RNA polymerase. In this study,
                      we developed an isopropyl-β-d-1-thiogalactopyranosid
                      (IPTG)-inducible T7 expression system in the prophage-free
                      strain C. glutamicum MB001. For this purpose, part of
                      the DE3 region of Escherichia coli BL21(DE3) including the
                      T7 RNA polymerase gene 1 under control of the lacUV5
                      promoter was integrated into the chromosome, resulting in
                      strain MB001(DE3). Furthermore, the expression vector pMKEx2
                      was constructed allowing cloning of target genes under the
                      control of the T7lac promoter. The properties of the system
                      were evaluated using eyfp as heterologous target gene.
                      Without induction, the system was tightly repressed,
                      resulting in a very low specific eYFP fluorescence
                      (= fluorescence per cell density). After maximal induction
                      with IPTG, the specific fluorescence increased 450-fold
                      compared with the uninduced state and was about 3.5 times
                      higher than in control strains expressing eyfp under control
                      of the IPTG-induced tac promoter with the endogenous RNA
                      polymerase. Flow cytometry revealed that T7-based eyfp
                      expression resulted in a highly uniform population, with
                      $99\%$ of all cells showing high fluorescence. Besides eyfp,
                      the functionality of the corynebacterial T7 expression
                      system was also successfully demonstrated by overexpression
                      of the C. glutamicum pyk gene for pyruvate kinase, which
                      led to an increase of the specific activity from 2.6 to
                      135 U mg−1. It thus presents an efficient new tool for
                      protein overproduction, metabolic engineering and synthetic
                      biology approaches with C. glutamicum.},
      cin          = {IBG-1},
      ddc          = {570},
      cid          = {I:(DE-Juel1)IBG-1-20101118},
      pnm          = {581 - Biotechnology (POF3-581)},
      pid          = {G:(DE-HGF)POF3-581},
      typ          = {PUB:(DE-HGF)16},
      UT           = {WOS:000350345300006},
      pubmed       = {25488698},
      doi          = {10.1111/1751-7915.12236},
      url          = {https://juser.fz-juelich.de/record/173351},
}