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The construction and application of a lux-based nitrate biosensor
A G Prest1, M K Winson, J R Hammond
1Bass Technical Centre, Burton on Trent, UK.
Letters in Applied Microbiology
|May 1, 1997
Summary
A novel Escherichia coli biosensor utilizing the nitrate reductase (narG) promoter and lux operon detects low nitrate levels. This tool also identifies bacterial regulatory protein NarL, aiding in brewery contaminant analysis.
Area of Science:
- Microbiology
- Molecular Biology
- Biosensor Technology
Background:
- Nitrate contamination is a concern in various industries, including brewing.
- Understanding nitrate reductase regulation is crucial for microbial analysis.
- Escherichia coli possesses a well-characterized nitrate reductase system.
Purpose of the Study:
- To develop a bioluminescent biosensor for nitrate detection in E. coli.
- To utilize the narG promoter-lux operon fusion for sensing.
- To investigate the presence of functional NarL regulatory protein in other bacteria.
Main Methods:
- Constructing a plasmid with a transcriptional fusion of the E. coli narG promoter and Photorhabdus luminescens lux operon.
- Introducing the plasmid into E. coli to create a bioluminescent biosensor.
- Testing the biosensor's sensitivity to nitrate.
- Using the biosensor to analyze the brewery contaminant Obesumbacterium proteus.
Main Results:
- The E. coli biosensor exhibited high bioluminescence in the presence of nitrate.
- The biosensor detected nitrate down to 5 x 10(-5) mol L-1 (0.3 ppm).
- Obesumbacterium proteus did not show nitrate-dependent bioluminescence, indicating regulatory differences.
Conclusions:
- A sensitive E. coli-based biosensor for nitrate detection was successfully developed.
- The biosensor can be used to infer the presence of functional NarL regulatory protein.
- This approach highlights regulatory divergence in nitrate reductase mechanisms among bacterial species.