coli strain was calculated from growth curves performed in LB check details medium at 37°C with chloramphenicol [Cm] 100 μg/ml or with spectinomycin [Sp] 100 μg/ml. The efficacy of propagation of the hybrid phage λimm P22  was measured on different strains. Table 3 presents the relative efficiency of plating (EOP) of each strain in comparison with that of the wild type parental strain. Phage
propagation on strain MG1655 ΔsmpB containing the empty vector pILL2150 was, as expected, strongly affected with an EOP of 1.3 × 10-5 (Table 3). Relative EOP of strain MG1655 ΔsmpB pILL786 in the presence of IPTG, expressing Hp-SmpB is close to 1 (Table 3). This result demonstrated that Hp-SmpB is active in E. coli and efficiently complemented the phage
propagation defect phenotype. In addition, the growth defect of MG1655 ΔsmpB mutant was analyzed with or without Hp-SmpB. Under our test conditions, MG1655 ΔsmpB mutant click here presented a doubling time that was about twice that of the wild type strain and was restored to wild type growth in the presence of Hp-SmpB expressed by pILL786 (Figure 2 and Table 3). This indicated that Hp-SmpB is able to replace Gefitinib cost Ec-SmpB functions during trans-translation
in E. coli. Figure 2 Doubling time of E. coli ΔssrA or ΔsmpB mutants expressing SmpB Hp WT, SsrA Hp WT or mutants. Doubling times were calculated for E. coli strains expressing SmpB Hp , SsrA Hp and different mutant versions of SsrA Hp from plasmids. Doubling times (g values) correspond to the mean generation time. As a selleck inhibitor control, growth complementation of the E. coli ΔssrA with Ec-ssrA is presented. Empty vector corresponds to a vector without insert. Table 3 Ability of H. pylori SmpB and of wild type or mutant alleles of ssrA Hp to support growth of λimm P22 in E. coli ΔssrA or ΔsmpB deletion mutants and to restore the growth defect in E. coli ΔssrA or ΔsmpB mutants Strains ssrA or smpB alleles EOP§ Growth defect restoration in E. coli ΔsmpB or in E. coli ΔssrA MG1655 smpB Ec ssrA Ec 1 – MG1655 ΔsmpB pILL2150 ΔsmpB Ec ssrA Ec 1.3 × 10-5 no MG1655 ΔsmpB pILL786 ΔsmpB Ec ssrA Ec /smpB Hp 0.6 yes MG1655 ΔssrA pILL2150 smpB Ec ΔssrA Ec 2.