Highly Resolved Detection of Long Non-coding RNAs In Situ

Abstract

Long non-coding RNAs (lncRNAs) have been postulated to function in a number of DNA-based processes, most notably transcription. The detection of lncRNAs in situ can offer insights into their function. Fluorescence in situ hybridization (FISH) enables the detection of specific nucleic acid sequences, including lncRNAs, within individual cells. Current RNA FISH techniques can inform both the localization and expression level of RNA transcripts. Together with advances in microscopy, these in situ techniques now allow for visualization and quantification of even lowly expressed or unstable lncRNAs. When combined with detection of associated proteins and chromatin modifications by immunofluorescence, RNA FISH can lend essential insights into lncRNA function. Here, we describe an integrated set of protocols to detect, individually or in combination, specific RNAs, DNAs, proteins, and histone modifications in single cells at high sensitivity using conventional fluorescence microscopy.