FIG 3.
Measuring cAMP levels in strains carrying mutations in pilT. (A) Graph depicting the average PPaQaYFP/PrpoDmKate per cell of selected strains during the first 6 h of surface attachment in glass well dishes, as described in the text and Materials and Methods. Solid lines represent the mean fluorescence of YFP/mKate per cell, and the shaded region represents the 95% confidence interval. At least 3 biological replicates were performed for each strain. A corresponding plot for all the pilT alleles described in this article can be found in Fig. S4. (B) Cells were grown on an agar surface for 5 h as described in Materials and Methods and then analyzed by flow cytometer to quantify the amount of intracellular cAMP via the PaQa reporter. These values were then normalized by the WT value for that biological replicate. Bars and errors bars represent the mean and standard deviation from 6 biological replicates. Strains that were able to twitch at >75% of the WT level are purple bars, strains that twitch at between 25 and 75% of the WT level are blue bars, and strains that twitch at <25% of the WT level are green bars. Data were analyzed by one-way ANOVA, followed by Tukey’s posttest comparison. *, P < 0.05; **, P ≤ 0.01; ****, P ≤ 0.0001.