Selection of phage-resistant mutants and genome analysis.

Host strains resistant to phage Bp7 were isolated during the phage cultivation process. Then, 100 μl E. coli K-12 (10⁹ CFU/ml) and 100 μl Bp7 (10⁹ PFU/ml) were mixed in 5 ml LB and cultured at 37°C overnight. After the culture turbidity changed from turbid to clear and became turbid again, 100 μl resistant strain culture was spread onto double-layer phage plates, and the lawn on the agar layer contained 10⁹ PFU/ml phage Bp7. The plates were incubated overnight, and colonies of phage-resistant mutants were purified by repeated single-colony isolation. A resistant strain named E. coli K-12-R was selected and sequenced by Biozeron Biotech Co., Ltd. (Shanghai, China). The genome sequence of the resistant strain was compared with that of the wild-type strain and analyzed to identify mutations that corresponded to phage resistance. The adsorption kinetics of phage Bp7 to E. coli K-12-R were assayed by the double-layer agar method, as described above.