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Open Access Highly Accessed Methodology article

Genetic tools for the investigation of Roseobacter clade bacteria

Tanja Piekarski1, Ina Buchholz2, Thomas Drepper3, Max Schobert1, Irene Wagner-Doebler2, Petra Tielen1* and Dieter Jahn1

Author Affiliations

1 Institute of Microbiology, Universität Braunschweig, Spielmannstrasse 7, D-38106 Braunschweig, Germany

2 Department for Cell Biology, Research Group Microbial Communication, Helmholtz Center for Infection Research, Inhoffenstrasse 7, D-38124 Braunschweig, Germany

3 Institute of Molecular Enzyme Technology, Heinrich-Heine- University Düsseldorf, Research Center Jülich, Stetternicher Forst, D-52426 Jülich, Germany

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BMC Microbiology 2009, 9:265  doi:10.1186/1471-2180-9-265

Published: 18 December 2009

Abstract

Background

The Roseobacter clade represents one of the most abundant, metabolically versatile and ecologically important bacterial groups found in marine habitats. A detailed molecular investigation of the regulatory and metabolic networks of these organisms is currently limited for many strains by missing suitable genetic tools.

Results

Conjugation and electroporation methods for the efficient and stable genetic transformation of selected Roseobacter clade bacteria including Dinoroseobacter shibae, Oceanibulbus indolifex, Phaeobacter gallaeciensis, Phaeobacter inhibens, Roseobacter denitrificans and Roseobacter litoralis were tested. For this purpose an antibiotic resistance screening was performed and suitable genetic markers were selected. Based on these transformation protocols stably maintained plasmids were identified. A plasmid encoded oxygen-independent fluorescent system was established using the flavin mononucleotide-based fluorescent protein FbFP. Finally, a chromosomal gene knockout strategy was successfully employed for the inactivation of the anaerobic metabolism regulatory gene dnr from D. shibae DFL12T.

Conclusion

A genetic toolbox for members of the Roseobacter clade was established. This provides a solid methodical basis for the detailed elucidation of gene regulatory and metabolic networks underlying the ecological success of this group of marine bacteria.