Figure 2.

Heteroduplex production in λRF953-infected AC227[pMB4] cells. Equal amounts (15 μg/lane) of DNA from clear lysates (B) or total DNA preparations (20 μg/lane) (C) of samples taken at the indicated time after infection were digested by PvuII and NdeI restriction endonucleases and subjected to PAGE and Southern blot hybridization as described in text. Artificial complementary heteroduplexes (st.) were prepared as described (27) and identified as described in Materials and Methods. Digested samples from infected recBC and recA mutants are presented as negative controls. The schematic diagram (A) illustrates the linear substrate, released by in vivo restriction of the infecting phage. The locations of the Chi octamers (χ), relevant restriction sites, and the BglII linker (triangles), inserted as a heteroallelic marker at the XmnI site, are indicated. The expected circular and linear products of recombination, initiated by invasion of the 3′ (left)- or 5′ (right)-ending strands of the lower homolog (see Fig. 1) and mismatches on the circular products are indicated. The locations of the electrophoretic bands of the homoduplexes (Homodup.) and the heteroduplexes made by pairing the strands ending 3′ (3′ het.) or 5′ (5′ het.) at the EcoRI-induced breaks are indicated. The kinetics of 3′ (□) and 5′ (▪) heteroduplex production in total cellular DNA preparations (D) was determined by phosphorimaging analysis of the Southern blot presented in C.