Isolation of a Specific Transcript Using Biotinylated Antisense Probes

RNA molecules play a central role in the regulation of gene expression, interacting with proteins and undergoing chemical modifications that affect their stability, location, and function. Studying these processes at the level of individual transcripts requires methods that allow selective isolation of RNA with minimal bias. While global RNA enrichment techniques provide a broad overview of the transcriptome, they often lack the resolution required to dissect transcript-specific interactions and modifications. Here we describe a hybridization-based protocol that uses biotinylated antisense DNA probes for selective transcript isolation. This approach can be used as a targeted strategy to enrich for specific RNA molecules, serve as a basis for studies of transcript-associated RNA-binding proteins (RBPome) or RNA modifications, and provide a refined perspective on post-transcriptional regulation. The protocol is outlined in detail, with a focus on the critical steps of probe design, hybridization, and purification, to ensure the highest degree of specificity and reproducibility in RNA capture experiments.