Figure 1.

Construction and characterization of orthogonal temperature-responsive biosensors. (A) Reversible thermal regulation based on CI⁸⁵⁷ that is sensitive to high temperature, and the output is a reporter GFP under the temperature-responsive promoter. Dimmer CI⁸⁵⁷ complex could bind to specific sequences OR when the temperature is low and block the binding of RNA polymerase to the promoter that is necessary for transcription initiation. When the temperature is high, the DNA-binding activity of CI⁸⁵⁷ is antagonized, and RNA polymerase can bind to the promoter initiating transcription. (B) Schematic diagram of hybrid promoters, which were constructed by integrating the CI⁸⁵⁷-binding site (OR₁ and OR₂) into different positions of promoter P_veg. The promoter core region ‘−35’, ‘−10’ and transcription initiation site ‘+1’ are annotated. (C) The activity of each hybrid promoter in strains with CI⁸⁵⁷ was tested at the different levels of temperature. The transition point was defined as the temperature at which the relative fluorescence intensity exceeds 150. (D) The fitting of temperature and the relative fluorescence intensity of promoter P_veg5. (E) The strength of hybrid promoter P_veg5 in strains with or without CI⁸⁵⁷, respectively. (F) The activity of temperature-responsive promoters with different transition points. GFP, green fluorescent protein; RNAP, RNA polymerase; OR, operator site. Data are presented as mean ± s.d. of three replicates.