Single Molecule Tracking: Visualizing Transcription of Endogenous Loci in Living Cells.

Live-cell imaging of RNA provides mechanistic insights into the spatiotemporal organization and regulation of transcription. The most widely used method for visualizing transcription in living cells is the MS2 aptamer system. During nascent RNA synthesis of an MS2-labeled gene, a transcribed array of MS2 RNA stem-loops is bound by fluorescently labeled MS2 coat proteins to enable the visualization and tracking of single RNA molecules. Here, we present a protocol for visualizing nascent RNA transcription at an endogenous gene locus. Given the importance of the native chromatin context in regulating transcription, using this approach is crucial for uncovering mechanisms and understanding gene expression dynamics.