Fig. 1
From: Dynamic and tunable metabolite control for robust minimal-equipment assessment of serum zinc
Dual-input promoters decouple cell growth and pigment production. a Design of a dual-input system that decouples pigment production from cell growth. Colorless cells are added to fresh medium or to a serum sample at a density high enough that cells are visible, and a small-molecule inducer is added to activate the color-response circuit. After a short incubation, different pigments (either violacein, lycopene, or β-carotene) are produced to indicate different zinc concentrations. b Circuit diagram and schematic depicting the design of a dual-input promoter that regulates production of the violacein pathway genes, using IPTG/LacI-regulation as an example. Violacein should only be produced in low zinc conditions when IPTG is present. Analogous schematics for AraC- and T7 RNAP- mediated control are in Supplementary Fig. 1. c Promoter architecture for the library of synthetic promoters. The −10 and −35 σ70 binding domains are marked, and lac and zur operator sites are shown in yellow and blue, respectively. For the PBadZnu promoter, the −10 binding domain is not explicitly marked because it is embedded in the Zur operator site, and the green box indicates the entire PBad promoter sequence that is upstream of the −35 binding domain. For PT7 variants, the consensus T7 RNAP binding domain is marked in gray. Supplementary Table 1 contains annotated promoter sequences for all constructed promoters. d Fluorescent characterization of all engineered promoters. Ideally, eGFP should only be produced in the + inducer/−zinc state, as indicated by example output from a hypothetical ideal promoter (Pideal). Each box in the heat map is the average of three biological replicates. Supplementary Fig. 2 shows averages with standard deviations. e Violacein production from the best-responding hybrid promoters of each group. A starter culture (−inducer/−zinc) was used to inoculate cultures that contained the appropriate inducer and different concentrations of zinc. Bars represent the average of three biological replicates, which are depicted as overlaying points. Images below the graph show representative cell pellets from each condition. Source data for d and e are provided in the Source Data file.
