Quantification of flhDCSm promoter activity and flagellin synthesis in S. marcescens CH-1 and CH-1ΔA at 30 and 37°Ca

Strain and tempP1 P2 Flagellin synthesis
    30°C1,351 ± 511.962,539 ± 1052.3160 ± 22.61
    37°C688 ± 311,099 ± 4323 ± 4
    30°C1,444 ± 301.143,271 ± 1731.1587 ± 41.12
    37°C1,268 ± 192,847 ± 18578 ± 9
  • a RNA was isolated from S. marcescens CH-1 and the rssA mutant strain (CH-1ΔA) grown on swarming plates for 2 h. flhDCSm P1 and P2 promoter activities were evaluated by primer extension using a FAM-labeled primer followed by electrophoresis on an ABI Prism 3100 capillary DNA genetic analyzer. The promoter activity was scored for relative fluorescence intensity by using the peak area with GeneMapper software, version 3.5. The amount of flagellin was evaluated by calculation of the relative intensities of Western blotting results using Scion Imaging software after incubation for 3 h on seeding plates. The promoter activities and amounts of flagellin are presented as arbitrary units. Each result is the mean ± standard error from three independent experiments, with triplicate samples in each assay. Ratios were calculated as the promoter activity or flagellin amount of a strain incubated at 30°C divided by that at 37°C.