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Expression and quantification of firefly luciferase under control ofRhizobium meliloti symbiotic promoters

✍ Scribed by Cebolla, Angel ;Ruiz-Berraquero, Francisco ;Palomares, Antonio J.

Publisher: John Wiley and Sons
Year: 1991
Weight: 542 KB
Volume: 6
Category: Article
ISSN: 0884-3996
DOI: 10.1002/bio.1170060307

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✦ Synopsis

We have tested the use o f firefly luciferase for monitoring regulated symbiotic nitrogen fixation gene expression. Broad-host-range plasmids carrying translational fusions o f Rhizobium melilotinifH, fixA and nifA promoters were constructed. Despite l o w levels o f promoter activity the absence o f Escherichia c o l i endogenous luminescence and the high sensitivity o f the bioluminescent assay for firefly luciferase allowed rapid screening for functional luciferase expression. Plasmids containing symbiotic promoter-luc fusions were established in R. meliloti. Luciferase activity was detected and measured in both vegetative and symbiotic cells giving comparable results with those obtained by beta-galactosidase assays. In addition, the luciferase assay was quicker, more sensitive and could be carried out with unrestricted cells. Furthermore, bioluminescence was high enough in alfalfa nodules containing nifH-luc fusion t o be observed by a dark-adapted eye and photographed.

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