Construction and Characterization of a Lux-Marked Phenanthrene Degrading Bacterium

Document Type: Research Paper

Authors

1 Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, I.R. Iran.

2 Department of Molecular and Cell Biology, University of Aberdeen, Institute of Medical Sciences, Foresterhill, Aberdeen AB25 2ZD, UK.

Abstract

Construction of a luminescent microbial biosensor as a sensitive and rapid biotechnological tool for monitoring
survival and activity of genetically engineered microorganisms (GEMs) in the contaminated environment has
been the main focus of this study. Four rifampicin resistant phenanthrene-degrading bacteria viz.,
Commamonas testosteroni GZ38A, C. testosteroni GZ39, Pseudomonas putida GZ44 and P. stutzeri P16
were made using gradient plate techniques. All resistant strains were transformed with the plasmid harboring
the mini-Tn5-tet transposon cassette, containing the luxAB genes to confer stability. C. testosteroni
GZ38A and P. stutzeri P16 were successfully luxmarked in this manner. It was found that lux-marked
strains of C. testosteroni GZ38A were not able to degrade phenanthrene. Although the maximum growth
rate of lux-marked strain was significantly lower (P < 0.05) than that of the wild type P. stutzeri P16, the P.
stutzeri P16-luxAB4 was selected because it has the following criteria: the stability of expression of tetresistance over 180 generations, phenanthrene - degradability and high level of luminescence light output
in the absence and presence of phenanthrene. The addition of 1 litter (0.5 % v/v) n-decyl aldehyde
produced consistently high levels of luminescence at various stages of selected strain. Results clearly indicated
that lux-marked strain was appropriately constructed and it is a novel biodegradative luminescent
biosensor which enables us to monitor the fate of a phenathrene bacterial degrader genetically or nongenetically made within a polluted environment.

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