Prophage excision and replication.

<p>Agarose gel analysis of prophage excision and circularization products corresponding to <i>attB</i> and <i>attP</i> regions, respectively, probed by PCR. (A) Experimental approach: two sets of primers were used to detect prophage excision from the chromosome. The first set in red targets the excision site on the chromosome (<i>attB</i>) and the second set in green targets prophage circular forms (<i>attP</i>). (B) Prophage excision products corresponding to <i>attB</i> region were probed by PCR in WT cultures induced with 2 µg/ml of mitomycin C (M), or ciprofloxacin (C) or uninduced (−) at 28, 37 and 42°C. Ch corresponds to amplification of a strain specific chromosomal gene. (C) Prophage excision products in strain <i>pp3⁻ pp5⁻</i> at 37°C. (D–F) Excision and circularization products probed by semi-quantitative PCR on 100, 10 and 1 pg of total bacterial DNA prepared from cultures of WT (D) and strains <i>pp3⁻ pp5⁻</i> (E) <i>pp4⁺</i> (F) induced for 2 h with 2 µg/ml of ciprofloxacin at 37°C except for detection of pp4 products in the WT strain that were obtained from DNA prepared after induction at 42°C. Twenty and 32 PCR cycles were used to amplify products of pp1 and pp7 and products of pp3, pp4 and pp5, respectively. These results are representative of three independent experiments.