Construction of a ColD cda promoter-based SOS-green fluorescent protein whole-cell biosensor with higher sensitivity toward genotoxic compounds than constructs based on recA, umuDC, or sulA promoters.
Four different green fluorescent protein (GFP)-based whole-cell biosensors were created based on the DNA damage inducible SOS response of Escherichia coli in order to evaluate the sensitivity of individual SOS promoters toward genotoxic substances. Treatment with the known carcinogen N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) revealed that the promoter for the ColD plasmid-borne cda gene had responses 12, 5, and 3 times greater than the recA, sulA, and umuDC promoters, respectively, and also considerably higher sensitivity. Furthermore, we showed that when the SOS-GFP construct was introduced into an E. coli host deficient in the tolC gene, the minimal detection limits toward mitomycin C, MNNG, nalidixic acid, and formaldehyde were lowered to 9.1 nM, 0.16 microM, 1.1 microM, and 141 microM, respectively, which were two to six times lower than those in the wild-type strain. This study thus presents a new SOS-GFP whole-cell biosensor which is not only able to detect minute levels of genotoxins but, due to its use of the green fluorescent protein, also a reporter system which should be applicable in high-throughput screening assays as well as a wide variety of in situ detection studies.
Norman
Anders
A
Department of Microbiology, University of Copenhagen, Sølvgade 83H, 1307 Copenhagen K, Denmark. hestbjerg@bi.ku.dk
Hestbjerg Hansen
Lars
L
Sørensen
Søren J
SJ
D001483:Base Sequence; D015374:Biosensing Techniques; D004259:DNA-Directed DNA Polymerase; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics; D008769:Methylnitronitrosoguanidine / Q000633:toxicity; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011938:Rec A Recombinases / Q000235:genetics*; D013014:SOS Response, Genetics
D029968:Escherichia coli Proteins; C053139:sulA protein, E coli; D008769:Methylnitronitrosoguanidine; D049452:Green Fluorescent Proteins; C078202:UmuC protein, E coli; D011938:Rec A Recombinases; D004259:DNA-Directed DNA Polymerase; C055218:UmuD protein, E coli