Stimulation of Recombination between Homologous Sequences on Plasmid DNA and Chromosomal DNA in Escherichia coli by N-acetoxy-2-acetylaminofluorene

• Published in: Proceedings of the National Academy of Sciences - PNAS 1984-05, Vol.81 (9), p.2831-2835
• Main Authors: Luisi-DeLuca, Cynthia, Porter, Ronald D., Taylor, William D.
• Format: Article
• Language: English
• Subjects: 2-Acetylaminofluorene - analogs & derivatives; Acetoxyacetylaminofluorene - pharmacology; Adducts; DNA; DNA Repair; DNA, Bacterial - genetics; Enzymes; Escherichia coli - genetics; Gels; Genetic mutation; Insect colonies; Lac operon; Molecules; Mutagenesis; Mutation - drug effects; Plasmids; Rec A Recombinases - genetics; Recombination, Genetic - drug effects
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.81.9.2831

A plasmid containing a wild-type lac operon and a tetracycline-resistance gene was covalently modified by N-acetoxy-2-acetylaminofluorene and used to transform two series of Lac-Escherichia coli cell types. Each set contained wild-type and repair-deficient mutants. One set of cells contained a lacY mutation and the other a deletion of the entire lac operon. Survival and mutagenesis of the plasmid were measured as a function of the N-acetoxy-2-acetylaminofluorene concentration. The results indicate that when no homologous sequences are present in the chromosomal DNA, mutations occur at a low frequency: at 10% survival the frequency was 1-2× 10-4mutants per transformant. When homologous sequences, the lacY allele, are present in the chromosomal DNA, Lac-plasmids are found at a high frequency in a recA-dependent, lexA-independent fashion: at 10% survival the frequency was 5-10× 10-2mutants per transformant. Southern blot analysis of the restriction enzyme profiles of the resulting plasmid and host-cell DNA sequences showed recombinational transfer of host sequences to the N-acetoxy-2-acetylaminofluorene-treated plasmid had occurred. When the host chromosomes contained Lac+homologous sequences no mutants were found, indicating that the results were not caused by error-prone recombination.