Incompatibility of DFHBI based fluorescent RNA aptamers with particular commercial cell-free expression systems

Fluorescent RNA aptamers are an increasingly used tool for quantifying transcription and for visualising RNA interactions, both in vitro and in vivo. However when tested in the commercially available, E. coli extract based Expressway cell-free expression system, no fluorescence is detected. The same experimental setup is shown to successfully produce fluorescent RNA aptamers when tested in another buffer designed for in vitro transcription, and RNA purification of the Expressway reaction products show that transcription does occur, but does not result in a fluorescent product. In this paper we demonstrate the incompatibility of a narrow selection of RNA aptamers in one particular cell-free expression system, and consider that similar issues may arise with other cell-free expression systems, RNA aptamers, and their corresponding fluorophores.