Abstract
AbstractFluorescent probes have been used for decades to illuminate RNA dynamics in cells and transparent organisms. Serial imaging remains challenging, though, owing to toxicities associated with the required excitation light. Applications in live animals are also limited due to difficulties in delivering external light to target tissues. To circumvent these issues, we developed an alternative method for visualizing RNAs that relies on bioluminescence. Bioluminescence involves a chemical reaction between luciferase enzymes and luciferin small molecules that produces photons. Since no excitation light is required, bioluminescent probes do not exhibit phototoxicity or photobleaching effects. Additionally, little to no background signal is produced, providing high signal-to-noise ratios and enabling sensitive detection in vivo – an important consideration for visualizing low-copy transcripts. We engineered a unique RNA sequence that recruits bioluminescent molecules upon transcription. We further optimized this system to modularly tag and visualize RNAs in a variety of contexts. These results provide the foundation for visualizing RNA dynamics in vivo.
Publisher
Cold Spring Harbor Laboratory