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Applied and Environmental Microbiology, October 2009, p. 6553-6558, Vol. 75, No. 20
0099-2240/09/$08.00+0     doi:10.1128/AEM.02825-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Development of a Single-Plasmid-Based Regulatable Gene Expression System for Borrelia burgdorferi{triangledown}

Christine R. Whetstine,{dagger} Joyce G. Slusser, and Wolfram R. Zückert*

Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, Kansas 66160

Received 12 December 2008/ Accepted 14 August 2009

We developed a single-plasmid-based regulatable protein expression system for Borrelia burgdorferi. Expression of a target gene is driven by Post, a hybrid B. burgdorferi ospA-tetO promoter, from a recombinant B. burgdorferi plasmid constitutively expressing TetR. The system was tested using the green fluorescent protein (GFP) as a reporter. Under noninducing conditions, recombinant B. burgdorferi cells were nonfluorescent, no GFP protein was detected, and residual, small amounts of transcript were detectable only by reverse transcription-PCR but not by Northern blot hybridization. Upon induction with anhydrotetracycline, increasing levels of GFP transcript, protein, and fluorescence were observed. This tight and titratable promoter system will be invaluable for the study of essential borrelial proteins. Since target protein, operator, and repressor are carried by a single plasmid, the system's application is independent of a particular strain background.

* Corresponding author. Mailing address: Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Mail Stop 3019, 3025 Wahl Hall West, 3901 Rainbow Boulevard, Kansas City, KS 66160-7420. Phone: (913) 588-7061. Fax: (913) 588-7295. E-mail: wzueckert{at}kumc.edu

{triangledown} Published ahead of print on 21 August 2009.

{dagger} Present address: Stowers Institute for Medical Research, 1000 E. 50th Street, Kansas City, MO 64110.

Applied and Environmental Microbiology, October 2009, p. 6553-6558, Vol. 75, No. 20
0099-2240/09/$08.00+0     doi:10.1128/AEM.02825-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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