Figure 2.—

Assay for prophage replication initiation from oriλ. (A) The nonexcisable cryptic λ fragment inserted (short arrow) within the E. coli chromosome remains repressed at 30° where the prophage repressor is active. Shifting cells to ≥38° inactivates the CI857 repressor for λ prophage transcription and replication initiation from oriλ. Multiple λ bidirectional replication initiation events from oriλ generate the onion-skin replication structure drawn at right. (B) λDNA (thick solid line) fragment within the E. coli chromosome (open boxes), BstEII restriction sites within λ and the E. coli chromosome showing bands generated by cleavage (Hayes et al. 1990), and the region amplified to prepare a DNA probe. (C) Assay for replication initiation from oriλ upon shifting culture cells to 42° to induce the cryptic prophage. Cells with plasmid pCI⁺ are not derepressed for λ transcription or oriλ replication at 42°, permitting a comparison between any chromosomal increase for cells shifted for 1 hr at 42° with oriλ replication initiation arising from the derepressed cryptic prophage in Y836 cells without the plasmid that were shifted for 1 hr to 42°. (See text for discussion of survivor CFU at 42°.) (D) The influence of host recombination defects on λDNA synthesis initiated from oriλ.