Detection and quantification of single mRNA dynamics with the Riboglow fluorescent RNA tag

Labeling and tracking biomolecules with fluorescent probes on the single molecule level enables quantitative insights into their dynamics in living cells. We previously developed Riboglow, a platform to label RNAs in live mammalian cells, consisting of a short RNA tag and a small organic probe that increases fluorescence upon binding RNA. Here, we demonstrate that Riboglow is capable of detecting and tracking single RNA molecules. We benchmark RNA tracking by comparing results with the established MS2 RNA tagging system. To demonstrate versatility of Riboglow, we assay translation on the single molecule level, where the translated mRNA is tagged with Riboglow and the nascent polypeptide is labeled with a fluorescent antibody. The growing effort to investigate RNA biology on the single molecule level requires sophisticated and diverse fluorescent probes for multiplexed, multi-color labeling of biomolecules of interest, and we present Riboglow as a new member in this toolbox.