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A system for detection of genetic and epigenetic alterations in Escherichia coli induced by DNA-damaging agents.
Journal of Molecular Biology
Date de publication
In order to compare the genetic and epigenetic effects of genotoxic agents, we have constructed Escherichia coli K12 strains that allow the detection of mutagenesis, SOS induction (epigenetic effect) and genetic recombination in the same genetic background. The epigenetic effect was detected in a similar way to any genetic alteration, i.e. by counting altered clones (colonies), using a gene fusion system that responds to a temporary epigenetic effect by a stable, heritable switch. The gene fusion consists of the E. coli gal operon and a partially deleted prophage lambda, resulting in the gal operon coming under the control of the cI and cro genes. It allows the detection of SOS induction and forward mutagenesis in the cI gene. Even a temporary inactivation of the CI repressor in this particular system leads to a stable epigenetic switch transmitted to the cellular progeny, which can be detected as Gal+ (red) colonies. The genetic (mutational inactivation of gene cI) and epigenetic (proteolytic inactivation of the product of gene cI) mechanisms leading to gal expression can be distinguished. Genetic recombination between two heteroallelic lacZ genes, one located in the bacterial chromosome, the other on an F'lac plasmid, can be detected as Lac+ colonies. Radiation and several chemical mutagens show very different capacities in generating mutants, inductants and recombinants; therefore, a dose range of any physical or chemical agent generates a set of relative values for the generation of mutants, inductants and recombinants that are characteristic of the agent.
Aflatoxin B1, Aflatoxins/pharmacology, Antibiotics, Antineoplastic/pharmacology, Carcinogens/pharmacology, DNA Repair, Escherichia coli/drug effects, Escherichia coli/genetics, Ethyl Methanesulfonate/pharmacology, Galactokinase/metabolism, Gamma Rays, Methylnitronitrosoguanidine/pharmacology, Mitomycin, Mitomycins/pharmacology, Mutagenicity Tests, Mutagens, Recombination, Genetic, Ultraviolet Rays
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