Expanding the genetic tool box for Cupriavidus necator by a stabilized L-rhamnose inducible plasmid system
| Autor: | Dirk Holtmann, Thomas Krieg, Ina Huth, Andreas Pannek, Anne Sydow, Stéphane E. Guillouet |
|---|---|
| Přispěvatelé: | Industrial Biotechnology, Centro de Investigación y Asistencia en Tecnología y Diseño del Estado de Jalisco (CIATEJ), Consejo Nacional de Ciencia y Tecnología [Mexico] (CONACYT)-Consejo Nacional de Ciencia y Tecnología [Mexico] (CONACYT), Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés (LISBP), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Recherche Agronomique (INRA), BMBF [031A226], German Academic Exchange Service (DAAD) [57212057], Institut National de la Recherche Agronomique (INRA)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS) |
| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
[SDV.BIO]Life Sciences [q-bio]/Biotechnology Rhamnose Cupriavidus necator L-rhamnose inducible expression system rhaP(BAD) protéine fluorescente Heterologous Bioengineering plasmid stability Applied Microbiology and Biotechnology Microbiology Green fluorescent protein 03 medical and health sciences chemistry.chemical_compound Plasmid plasmid gram negative bacteria Escherichia coli partitioning system fluorescent protein Inducer [SPI.GPROC]Engineering Sciences [physics]/Chemical and Process Engineering Promoter Regions Genetic fermentation expression des protéines biology General Medicine Gene Expression Regulation Bacterial biology.organism_classification Fluorescence Cell biology 030104 developmental biology [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology chemistry Metabolic Engineering bactérie gram négatif plasmide Fermentation Biotechnology Plasmids |
| Zdroj: | Journal of Biotechnology Journal of Biotechnology, Elsevier, 2017, 263, pp.1-10. ⟨10.1016/j.jbiotec.2017.10.002⟩ Journal of Biotechnology, 2017, 263, pp.1-10. ⟨10.1016/j.jbiotec.2017.10.002⟩ |
| ISSN: | 0168-1656 |
| DOI: | 10.1016/j.jbiotec.2017.10.002⟩ |
| Popis: | The Gram negative bacterium Cupriavidus necator is well known for the accumulation of poly(3-hydroxybutyrate) and its fast lithoautotrophic growth, leading in high cell densities. Although the host was engineered for the heterologous production of diverse chemicals and biopolymers in recent years, tool box of stabilized inducible expression systems is still limited. To avoid plasmid loss during fermentation processes and to allow expression of complex proteins, a tunable L-rhamnose inducible system was established and characterized using enhanced green fluorescent protein (eGFP). The construct was stabilized by a previously established partitioning system. An increase of fluorescence signal intensity in different media was shown with inducer concentrations up to 11 mM L-rhamnose. The strongest effects were measured at quite low concentrations - high tunability was observed between 0 and 0.4-1 mM (depending on the medium used). Expression is tightly regulated and could be increased over 140-fold in complex medium and approximately 60-fold in minimal medium due to induction with 11 mM L-rhamnose. Varying induction times were characterized regarding growth behavior and expression pattern, taking into consideration problems that may arise during expression of toxic proteins. The novel plasmid expands the tool box for engineering the highly flexible production host C. necator. |
| Databáze: | OpenAIRE |
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