Figure 1.

Assay for the selection of oxidative damage-induced frameshift mutations in repetitive sequences. A modified version of the low copy number plasmid pBR322 was constructed to contain the tetracycline resistance gene and a carbenicillin resistance gene containing an out-of-frame microsatellite sequence. This construct renders bacteria tetracycline-resistant and carbenicillin-sensitive. Following treatment with H₂O₂, damaged plasmid was transformed into wild-type BL21 E. coli. Colony growth on tetracycline-containing plates was indicative of plasmid survival, whereas growth on tetracycline plus carbenicillin was indicative of a compensating frameshift mutation to restore the reading frame to generate functional β-lactamase. Mutation frequency is calculated as the number of carbenicillin-resistant mutants recovered per tetracycline-resistant survivor.