Synthetic logic circuits using RNA aptamer against T7 RNA polymerase
With the growing appreciation for RNA molecules as the key component of genetic regulatory networks, efforts to utilize the programmable structure and function of RNA molecules for synthetic biology have accelerated in the recent past. In this study, we constructed synthetic circuits with RNA aptamer against T7 RNA polymerase and demonstrated its capability as a modular regulatory component. Due to the wide?spread usage of T7 RNAP for in vitro and in vivo applications, the T7 RNAP aptamer can be a useful addition to the regulatory toolkit for synthetic biology and metabolic engineering.AbstractRecent advances in nucleic acids engineering introduced several RNA?based regulatory components for synthetic gene circuits, expanding the toolsets to engineer organisms. In this work, we designed genetic circuits implementing an RNA aptamer previously described to have the capability of binding to the T7 RNA polymerase and inhibiting its activity in vitro. We first demonstrated the utility of the RNA aptamer in combination with programmable synthetic transcription networks in vitro. As a step to quickly assess the feasibility of aptamer functions in vivo, we tested the aptamer and its sequence variants in the cell?free expression system, verifying the aptamer functionality in the cell?free testbed. The expression of aptamer in E. coli demonstrated control over GFP expression driven by T7 RNA polymerase, indicating its ability to serve as building blocks for logic circuits and transcriptional cascades. This work elucidates the potential of T7 RNA polymerase aptamer as regulators for synthetic biological circuits and metabolic engineering.