Abstract
RNA toehold switches are powerful tools that can be used as biosensors to detect nearly any RNA sequence. In the presence of a specific RNA trigger, the toehold switch allows translation of a reporter protein. Toehold switches expressed in cell-free expression systems have been used as biosensors for several viruses and bacterial RNAs. Here we explore the use of the NanoLuc complementation reporter NanoBiT and found that it provides improved sensitivity over sfGFP. This reporter can be applied broadly across other RNA toehold switch applications.