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Duplex DNA oligomer constructs (32 base pairs) were prepared that contained a single benzo[alpyrene (BP) adduct at a specific deoxyadenosine or deoxyguanosine site in either one or both strands. These constructs were inserted into M13 replicative form viral DNA, and the DNA from progeny virus generated by transfection of Escherichia coli was examined by sequence analysis at the site of oligomer insertion. With nonalkylated constructs, and with constructs containing only one BP adduct, no sequence alterations were found in progeny viral DNAs. With constructs containing two BP adducts, one in each strand and closely spaced, some progeny DNAs showed the original oligomer sequence, whereas others exhibited large deletions and illegitimate (nonhomologous) recombination, both of which removed the damaged construct. Increasing the distance between BP adducts in the construct reduced the frequency of recombinant events. These sequence alterations occurred in both recA + and recAhost cells. We speculate that the closely spaced adducts in opposite construct strands cause a rare distortion in DNA structure, which activates the recombinant machinery, and that mutagenic and carcinogenic agents other than polycyclic aromatic hydrocarbons may cause similar DNA distortions, which induce illegitimate recombination.In an earlier report (1), we described the use of short DNA oligomers as targets for alkylation by benzo[a]pyrene diol epoxide (BPDE), the activated metabolite of benzo[a]pyrene (BP) (2), for the study of mutagenesis in Escherichia coli. In our experimental system, E. coli was transfected with phage M13 replicative form (RF) DNA, which was modified to contain a 10-base-pair (bp) BamHI linker treated with BPDE. Sequence analysis of DNA isolated from progeny virus indicated the presence of large deletions at the site of oligomer insertion. It was speculated that the relatively high localized concentration of multiple BP adducts in the linker target might be responsible for the large deletions we observed.To examine this phenomenon in greater depth, we prepared several duplex oligomers having BP adducts at a single site in each DNA strand, but with different spacings between the two adducts. These site-specific constructs were inserted into M13 RF DNA, and, after transfection of host cells, progeny viruses were selected for sequence analysis at the DNA region of oligomer insertion. The results of these analyses are summarized as follows: some progeny viral DNAs contained the original oligomer insert sequence, whereas others were modified exhibiting large deletions and insertions of illegitimate (nonhomologous) recombinant segments, both of which removed the damaged construct. Similar DNA modifications were observed with constructs alkylated either at deoxyadenosine or deoxyguanosine residues, but increasing the distance between BP adducts in the construct reduced the frequency of recombinant events. The induction of deletions and recombinants occurred in both recA + and recA -host cells. MATERIALS AND METHODS

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