Survival and mutagenesis of bacterial plasmids with localized carcinogen adducts

Restriction fragments from nonessential gene regions in bacterial plasmids were covalently modified with benzo[a]pyrene-7,8-dihydrodiol-9,10-oxide (BPDE) and ligated to the remaining unmodified fragment to construct the plasmid with a localized patch of BPDE adducts. These preparations were introduced into appropriate strains of E. coli and the effect of the carcinogen adducts on plasmid survival and mutagenesis determined. Survival as a function of adduct concentration of randomly modified plasmids was the same as that of plasmids with localized adducts in both repair-proficient and -deficient strains, indicating that the simple presence of the carcinogen is the main factor in plasmid mortality. The results of these and other experiments indicate that plasmid survival is a function of the adduct/plasmid molecule ratio not the adduct/nucleotide ratio. Plasmids with mutations produced in the region (tetracycline resistance) containing the localized adducts were selected and the nature of the mutations determined by direct sequence analysis. The mutations included frameshifts, transitions, and transversions.