Long noncoding RNAs (lncRNAs) are a family of non-protein-coding RNA transcripts greater than 200 nucleotides in length that have been regarded as crucial modulators of gene expression in various biological and disease contexts, but mechanisms underlying such regulation still remains largely elusive. In addition to cell lysate-based approaches that have proven invaluable for studies of lncRNAs, live-imaging methods can add value by providing more in-depth information on lncRNA dynamics and localizations at the single-molecule level. Recently, we have developed a versatile imaging approach based on molecular beacons (MBs), which are a class of fluorogenic oligonucleotide-based probes with the capacity to convert RNA target hybridization into a measurable fluorescence signal. In this chapter, we describe the detailed protocol of using MBs to illuminate lncRNA transcripts at the single-molecule level in living cells.
Keywords: Molecular beacons; Single-molecule; Tandem repeats; lncRNA.