Fig. 6.
A regulated TX device is more robust than an unregulated device with comparable output level. a AHL dose responses of the regulated (black square in top panel) and unregulated (white circle in top panel) TX devices. The regulated device uses a stronger Ec-TTL-P109 promoter and the unregulated device uses a weaker BBa_J23116 promoter. For both circuits, the RBS TIRs of ECF32 are 6474, corresponding to high feedback gain. The robustness vs. nominal output chart of Fig. 4b is reported here to facilitate comparison. Error bars represent standard deviation from six replicates, including three biological replicates each having two technical replicates. Specific value of an independent experiment is presented as a black dot. b The mRNA level of mRFP1 gene was quantified by RT-qPCR and normalized to the reference gene cysG (see Method for details). Data represent mean values (±standard deviation). c Temporal responses of the unregulated and the regulated devices were monitored in parallel by microplate photometer. Cells were first grown in multiple batches in the absence of AHL for 8 h till they reach steady state GFP expression. Each batch was diluted every 2 h to maintain exponential cell growth. The resource competitor was then induced at t = 0 with the indicated AHL concentrations. Sampling time interval of microplate photometer was 2 min. Mean GFP per OD values in arbitrary unit (A.U.) are normalized to those of samples without AHL induction to reflect relative change in GFP expression. Error bars represent standard deviation from three biological replicates. Temporal data without normalization can be found in Supplementary Fig. 9. d GFP fluorescence histograms (normalized to the statistical mode) of three biological replicates measured by flow cytometry 6 h after AHL induction. Respective scatter plots of both RFP and GFP and their detailed statistics can be found in Supplementary Note 6. Data in a–d were collected from three independent experiments. Source data are provided as a Source Data file.