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Construction of a nrdA::luxCDABE Fusion and Its Use in Escherichia coli as a DNA Damage Biosensor

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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.

No MeSH data available.


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Maximum relative luminescence values seen from strain BBTNrdA after exposure to different concentration of (A) phenol, (B) 2-chlorophenol (2-CP), (C) 2,4-dichlorophenol (2,4-DCP) and (D) 2,4,5-trichlorophenol (2,4,5-TCP).
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f4-sensors-08-01297: Maximum relative luminescence values seen from strain BBTNrdA after exposure to different concentration of (A) phenol, (B) 2-chlorophenol (2-CP), (C) 2,4-dichlorophenol (2,4-DCP) and (D) 2,4,5-trichlorophenol (2,4,5-TCP).

Mentions: Furthermore, additional experiments were conducted using membrane-damaging chemicals, i.e., phenol, 2-chlorophenol (2-CP), 2,4-dichlorophenol (2,4-DCP), and 2,4,5-trichlorophenol (2,4,5-TCP) [34]. Figure 4 shows that there was no response to these chemicals. This was expected since these compounds should have no effect on the structure or replication of the cellular DNA. Taken together with the results from the oxidative compounds, these results demonstrate that the nrdA gene expression level is not induced by membrane damaging or oxidative toxicants, but only by DNA damaging compounds.


Construction of a nrdA::luxCDABE Fusion and Its Use in Escherichia coli as a DNA Damage Biosensor
Maximum relative luminescence values seen from strain BBTNrdA after exposure to different concentration of (A) phenol, (B) 2-chlorophenol (2-CP), (C) 2,4-dichlorophenol (2,4-DCP) and (D) 2,4,5-trichlorophenol (2,4,5-TCP).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3927504&req=5

f4-sensors-08-01297: Maximum relative luminescence values seen from strain BBTNrdA after exposure to different concentration of (A) phenol, (B) 2-chlorophenol (2-CP), (C) 2,4-dichlorophenol (2,4-DCP) and (D) 2,4,5-trichlorophenol (2,4,5-TCP).
Mentions: Furthermore, additional experiments were conducted using membrane-damaging chemicals, i.e., phenol, 2-chlorophenol (2-CP), 2,4-dichlorophenol (2,4-DCP), and 2,4,5-trichlorophenol (2,4,5-TCP) [34]. Figure 4 shows that there was no response to these chemicals. This was expected since these compounds should have no effect on the structure or replication of the cellular DNA. Taken together with the results from the oxidative compounds, these results demonstrate that the nrdA gene expression level is not induced by membrane damaging or oxidative toxicants, but only by DNA damaging compounds.

View Article: PubMed Central - PubMed

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.

No MeSH data available.


Related in: MedlinePlus