Table 1.

Mean ± SEM (standard errors of the mean) of apparent FRET efficiency for wild-type and ΔegcpBΔecpC cells expressing the c-di-GMP sensor YFP-YcgR₃₉₃₇-CFP.

Sample	12 hrs/0 μM	12 hrs/50 μM	12 hrs/100 μM	24 hrs/50 μM
Wild type	0.253 ± 0.002 (n = 10)	0.310 ± 0.002 (n = 11)	0.363 ± 0.001 (n = 5)	0.402 ± 0.003 (n = 10)
ΔegcpBΔecpC mutant	0.257 ± 0.003 (n = 10)	0.291 ± 0.002 (n = 10)	0.339 ± 0.004 (n = 5)	0.405 ± 0.002 (n = 10)

Various induction levels were tested (listed as ‘time μM⁻¹’ IPTG in the table) to establish the dynamic range of the sensor. The sensor was sensitive to changes in the concentrations of c-di-GMP when it was incubated for approximately 12 h with 50 to 100 μM IPTG. An order-of-magnitude estimate of the sensor concentration based on the intensity of donor emission corrected for FRET, F^D (Patowary et al., 2013), as described in the Experimental procedures section, suggested that the sensor expression level varied between roughly 10 molecules per cell, for 12 h incubation with 0 μM IPTG (first column), and 1000 sensor molecules per cell, for 24 h incubation with 50 μM IPTG (fourth column). The sensor concentration around which the sensor responded to changes in c-di-GMP concentrations, shown in the second and third columns in the Table, were on the order of 100 sensor molecules per cell. Within that concentration range, significant differences between the FRET efficiencies of the wild type and ΔegcpBΔecpC mutant were observed. n = number of FRET images. FRET images contained an average of 50 cells per image.