Abstract
The promoter of nrdA gene which is related with DNA synthesis was used to construct a DNA damage sensitive biosensor. A recombinant bioluminescent E. coli strain, BBTNrdA, harboring a plasmid with the nrdA promoter fused to the luxCDABE operon, was successfully constructed. Its response to various chemicals including genotoxic chemicals substantiates it as a DNA damage biosensor. In characterization, three different classes of toxicants were used: DNA damaging chemicals, oxidative stress chemicals, and phenolics. BBTNrdA only responded strongly to DNA damaging chemicals, such as nalidixic acid (NDA), mitomycin C (MMC), 1-methyl-1-nitroso-N-methylguanidine (MNNG), and 4-nitroquinoline N-oxide (4-NQO). In contrast, there were no responses from the oxidative stress chemicals and phenolics, except from hydrogen peroxide (H2O2) which is known to cause DNA damage indirectly. Therefore, the results of the study demonstrate that BBTNrdA can be used as a DNA damage biosensor.
Highlights
Due to environmental pollution, specific and sensitive detection methods are in need for environmental contaminants
SOS-dependent bacterial test systems is used for DNA-damaging agents, and their response for those chemicals is known as SOS response
Strain BBTNrdA was constructed by fusing the nrdA promoter with the luxCDABE operon in plasmid pDEW201 and transforming this plasmid into E. coli strain RFM443
Summary
Specific and sensitive detection methods are in need for environmental contaminants. The reaction time needed to generate the bioluminescent responses is very short Using this procedure, the recA, sulA, umuCD and recN promoters have previously been fused with the luxCDABE genes and the strains carrying these fusions have been used widely in toxicity assays [11,12,13,14]. In this study we developed BBTNrdA, a cell-based genotoxicity sensor which is specific in its responses to genotoxins This E. coli strain harbors a plasmid with the nrdA promoter fused to the luxCDABE operon. Characterization of this strain was performed using exposures to DNA damaging chemicals, oxidative stress-inducing chemicals and phenolics. The results clearly show that BBTNrdA strongly responded to only genotoxic compounds
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