Genetic design and construction of the synthetic dual operon. (a) Design principle. The mutually inhibitory circuit generally provides two stable states (red and green frames) for the cells. Only one fragment in the dual operon (red and green arrows) can be activated and achieves one of the two essential biological roles (red and green circles). (b) Genetic construction. The synthetic dual operon (in red and green) presenting two physiological roles (leucine and histidine biosynthesis, resp.) essential for cell growth was integrated into the E. coli genome. leuB and hisC expression were controlled by lacI and tetR, and reported by mCherry (red fluorescent protein) and gfpuv5 (green fluorescent protein), respectively. The genes lacI-Y, leuB and hisC within their native operons (Lac operon, Leu operon and His operon) were genetically disrupted.
