Molecular Technologies for Intracellular mRNA Imaging and Their Recent Biological Applications

Understanding the spatial and temporal behaviors of individual mRNA molecules is essential for elucidating gene expression regulation and cellular function. Imaging these processes within cells offers unique insights into transcriptional heterogeneity, mRNA transport, translation, and degradation dynamics. In this Review, we summarized the principal molecular technologies for intracellular mRNA imaging and tracking. These approaches are classified based on their compatibility with fixed cells versus live cells. Fixed-cell FISH-based methods provide spatial resolution and multiplex quantification. Live-cell systems, such as RNA stem-loop binding proteins, CRISPR/Cas platforms, fluorogenic RNA aptamers, fluorescent chemical labeling strategies, molecular beacons, and DNA circuits, enable real-time visualization of both endogenous and exogenous mRNA transcripts in live cells. We finally discuss the challenges and potential future directions for intracellular mRNA imaging. We anticipate that this Review will inspire the development of the next generation of mRNA imaging technologies in diverse biological applications.