Fig. 1. (A) Denaturing PAGE (8%) characterization for the preparation of Apt/Dz. Lane 1: Apt; lane 2: Apt + AS1 + ligase; lane 3: Dz; lane 4: Dz + AS2 + ligase; lane 5: circularized Apt + Dz + AS2 + ligase; lane 6: purified Apt/Dz. The concentrations of Apt, Dz, AS1, AS2 and ligase were 500 nM, 500 nM, 2 μM, 2 μM, and 40 U, respectively. Exonuclease I/III was used to degrade the un-ligated ssDNA. (B) Fluorescence assays for TK1 mRNA-induced activation of Dz activity in Apt/Dz: (a) S; (b) Apt/Dz + S; (c) Apt/Dz + S + TK1 mRNA-1; (d) Apt/Dz + S + TK1 mRNA-3; (e) Apt/Dz + S + TK1 mRNA; (f) Apt/Dz* + S + TK1 mRNA. The concentrations of S, Apt/Dz, Apt/Dz*, TK1 mRNA-1 and TK1 mRNA-3 were all 100 nM and the concentrations of the TK1 mRNAs were 50 nM. Solutions were incubated for 2 h in the presence of 10 mM Mg²⁺, followed by fluorescence measurements.