The plates were then incubated overnight at 28°C, and AHL is indicated by the presence of a blue spot. Western blotting analysis Bacterial cultures were grown in NYG medium overnight and inoculated in the same medium. The refreshed cultures were grown at 37°C to an OD600 of 4.5; and 1 ml of each bacterial culture was collected and centrifuged. The cells were lysed by adding 250 μl celLyticTM B cell Lysis Reagent (Sigma). The concentrations of total protein samples were measured and normalized. Then the samples were KPT-8602 denatured by boiling for 10 min and separated by 10% SDS-PAGE. Western blot analysis was performed following
the standard protocols [39]. Acknowledgements The funding for this work was provided by the Biomedical Research Council, the Agency of Science, Technology and Research (A*Star), Singapore. Electronic supplementary material
Additional file 1: Figure S1: Mutation of BCAM0227 does not affect cepI expression level. (DOC 26 KB) Additional file 2: Figure S2: Complementation of rpfR with RpfR, RpfRAAL and RpfRGGAAF. (DOC 28 KB) Additional file 3: Figure S3: Cumulative effect of BDSF and AHL systems in regulation of bacterial motility, biofilm formation, and protease production. (DOC 62 KB) Additional file 4: Table S1: Primers used in this study. (DOC 28 KB) References 1. selleck kinase inhibitor Federle MJ, Bassler BL: Interspecies communication in bacteria. J Clin Invest 2003, 112:1291–1299.PubMed 2. Whitehead NA, Barnard AM, Slater H, Simpson NJ, Salmond GP: Quorum-sensing in Gram-negative bacteria. FEMS Microbiol Rev 2001, 25:365–404.PubMedCrossRef
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