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When Escherichia coli K-12 (X) lysogens were infected with A phages, genetic exchanges between phage and prophage occurred at low frequencies (less than 0.1% between the markers P3 and P80), but at frequencies above 1% if the infecting phages were first treated with the photosensitizing agent 4,5',8trimethylpsoralen and 360 nm light. Exchanges were induced by psoralen damage at about the same frequency in wild-type lysogens and in those carrying recB-, recC-, recF-, or lexA -, but at an intermediate frequency in a quadruple mutant carrying recB recC-recF-sbcB-. Few if any exchanges were induced in lysogens carrying uvrA -, uvrB-, or recA-. The increase in the frequency of recombination was presumably due to the psoralen damage in the phage DNA molecules and the action of host cell repair and recombination enzymes. The production of crosslinks in the phage DNA by psoralen and 360.nm light was measured by sedimentation in alkali. It showed second-order kinetics indicative of a twophoton reaction. In contrast, first-order kinetics had been reported for monoadduct formation. Second-order kinetics, similar to those for crosslink production, were found for genetic exchanges in homoimniune crosses. Presumably, crosslinks, rather than monoadducts, cause most of the exchanges. Because the uvrA+ gene product (UV-endonuclease) was required, it is likely that recombination was initiated by DNA molecules cut at crosslinks. This system has been used to show that after the crosslinked phage duplex has been cut, one or more of the subsequent steps-homologous pairing, cutting, and joiningrequire the recA+ gene product. Studies on the mechanism of genetic recombination in various organisms indicate that recombinants can be formed by joining portions of two homologous molecules (1-3). It is supposed that DNA molecules from the two parents are first nicked, permitting single strands from each to pair in a heteroduplex. The hydrogen-bonded molecules are then enzymatically trimmed and joined to form recombinant molecules (4-8). Research into the underlying mechanisms has been hindered because of the low frequency with which recombination normally takes place and the difficulty of identifying suitable substrates for initiating recombination.One approach to this problem is to study systems in which the frequency of recombination is increased through the use of chemicals or radiation. Recombination is readily induced in microorganisms by ultraviolet-irradiation (9). The increased frequency is thought to be caused, in part, by the replication of the irradiated DNA and the formation of postreplication gaps opposite pyrimidine dimers (10-12). Recombination is also induced by agents that crosslink cellular DNA. In this regard, the efficiency of bifunctional agents is much greater than that of related monofunctional compounds (13).We chose to use Escherichia coli (A) lysogens infected with phage A for an investigation into genetic recombination induced by damage in phage DNA, because little if any replication of the damaged DNA would...

The effects of the mutation pairs recB21 recF143 and recB21 uvrD152 on the frequency of genetic recombination were investigated in A phage-prophage crosses under homoimmune conditions. To prevent recombinants from being formed by the phage red system, these experiments were performed with phages and prophages carrying red and gam mutations. Both spontaneous and damageinduced recombination was measured, the phages being either undamaged or treated with trimethylpsoralen and 360-nm light to cross-link the phage DNA. Control and damaged phages were allowed to infect lysogenic host cells under conditions in which phage gene expression was repressed and phage DNA replication was blocked by A immunity. Although the double mutations recB21 recF143 and recB21 uvrD152 reduced recombination in Hfr by F-crosses to 0.3 to 0.02% of the wild-type controls, the presence of these pairs of mutations in the host lysogens had relatively little effect on the results of the phage-prophage crosses. In the latter system, recB21 recF143 reduced spontaneous and damagedinduced recombination by less than threefold whereas recB21 uvrD152 increased it to three times the wild-type level, the increase being attributable to the uvrD mutation. Evidently, the gene products of recB,C, uvrD, and recF were not needed for A phage-prophage recombination under repressed conditions.

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