Single-Cell Imaging Approaches for Studying Small-RNA-Induced Gene Regulation.

RNA interference (RNAi) is a process by which gene expression is downregulated by small interfering RNAs or microRNAs. The quantification of the RNAi efficiency can be performed at both the messenger RNA (mRNA) and the protein level, which is required to assess the potency of small interfering RNAs or microRNAs. Recently, we employed a single-cell mRNA imaging method to study RNAi in which we visualized individual mRNA targets with high precision while resolving the cellular localization and cell-to-cell heterogeneity in addition to RNAi efficiency. In this Biophysical Perspective, we highlight our recent work on quantitative analysis of the RNAi pathway and point out some important future directions. Alongside, we discuss about several single-cell imaging techniques that can be applied to study RNAi. The single-cell imaging techniques discussed here are widely applicable to other gene regulation processes such as the CRISPR-CAS system.