Fig. 3: Remote toehold EP architecture uncouples tradeoff between sensitivity and dynamic range.

a Schematic of the remote toehold EP design, in which the polyA loop of Cbe pfl is replaced by a synthetic hairpin connected to the AD and invader by single stranded extendable polyA tethers. b Vfold3D model of a nucleated terminator featuring a remote toehold extension with a 5A 5’ tether and a 1A 3’ tether (5:1). c Dose responses of select remote toehold variants with varying length 5’ tethers, including the 5:1 design shown in (b). Vertical lines indicate EC₅₀ values of the curve with the corresponding color. d EC₅₀ measurements for remote toehold variants with varying length 5’ and 3’ tethers. n:1 indicates the length of the 5’ tether has been extended, with the length of the 3’ tether fixed at 1A. 1:m indicates the length of the 3’ tether has been extended, with the length of the 5’ tether fixed at 1A. n:m indicates the length of both tethers have been simultaneously extended. e Fold change measurements for variants in (d). f EC₅₀ and fold change data from (d) and (e) plotted against each other. Increasing opacity represents longer overall tether length. Data in panel (c) are dose response curves over n = 9 biological replicates, with points indicating mean and error bars indicating standard deviation. Data in panels (d), (e), and (f) are determined as described in Methods, from dose response curves including n = 9 biological replicates at each of 8 concentrations. Error bars in (d), (e), and (f) indicate standard error of the fit parameter. Source data are provided as a Source Data file.