A universal method for labeling native RNA in live bacterial cells

Abstract

The spectrum of RNA functions in the cell continues to widen and new types of RNA molecules continue to be discovered. However, methods to access and manipulate endogenous RNAs in live cells are limited. Here we describe a universal technique for labeling natural RNAs in live cells with the probes synthesized by the cell. The method is based on fluorescent protein complementation in combination with a split aptamer approach. Two RNA probes containing split aptamer sequences flanked with the antisense RNA target sequences are assembled on the target RNA to form a fluorescent ribonucleoprotein (RNP) complex. The mechanism of complex formation ensures highly sensitive RNA detection allowing visualization of endogenous bacterial mRNAs. We demonstrate the great potential of this method by detecting chromosomally low-level expressed unmodified bacterial mRNA in living bacterial cells. This method holds promise to become a broadly used tool in basic research, and eventually in diagnostics and therapeutics.