Detection of DNA damage by use of Escherichia coli carrying recA'::lux, uvrA'::lux, or alkA'::lux reporter plasmids.
Plasmids were constructed in which DNA damage-inducible promoters recA, uvrA, and alkA from Escherichia coli were fused to the Vibrio fischeri luxCDABE operon. Introduction of these plasmids into E. coli allowed the detection of a dose-dependent response to DNA-damaging agents, such as mitomycin and UV irradiation. Bioluminescence was measured in real time over extended periods. The fusion of the recA promoter to luxCDABE showed the most dramatic and sensitive responses. lexA dependence of the bioluminescent SOS response was demonstrated, confirming that this biosensor's reports were transmitted by the expected regulatory circuitry. Comparisons were made between luxCDABE and lacZ fusions to each promoter. It is suggested that the lux biosensors may have use in monitoring chemical, physical, and genotoxic agents as well as in further characterizing the mechanisms of DNA repair.
Vollmer
A C
AC
Swarthmore College, Pennsylvania 19081-1397, USA. avollme1@swarthmore.edu
Belkin
S
S
Smulski
D R
DR
Van Dyk
T K
TK
LaRossa
R A
RA
D000251:Adenosine Triphosphatases / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D015374:Biosensing Techniques; D004249:DNA Damage / Q000187:drug effects* / Q000528:radiation effects*; D045647:DNA Glycosylases; D004269:DNA, Bacterial / Q000187:drug effects* / Q000528:radiation effects*; D004268:DNA-Binding Proteins / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D004307:Dose-Response Relationship, Radiation; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D007763:Lac Operon; D049449:Luminescence; D008769:Methylnitronitrosoguanidine / Q000494:pharmacology; D008937:Mitomycins / Q000494:pharmacology; D008969:Molecular Sequence Data; D009153:Mutagens / Q000494:pharmacology; D009699:N-Glycosyl Hydrolases / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D011938:Rec A Recombinases / Q000235:genetics; D011995:Recombination, Genetic; D012097:Repressor Proteins; D015534:Trans-Activators; D014466:Ultraviolet Rays
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D008937:Mitomycins; D009153:Mutagens; D012097:Repressor Proteins; D015534:Trans-Activators; C064778:LuxR autoinducer binding proteins; D008769:Methylnitronitrosoguanidine; D011938:Rec A Recombinases; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C076131:DNA-3-methyladenine glycosidase II; C068631:UvrA protein, E coli; D000251:Adenosine Triphosphatases