Abstract
Mammalian cells express hundreds of RNA binding proteins (RBPs) that are essential regulators of RNA metabolism. RBP activity plays a central role in the control of gene expression programs and identification of RNA-protein interactions is critical for comprehensive understanding of gene regulation in cells. In recent years, various tools and techniques to identify these RNA-protein interactions have been developed. Among those, RNA immunoprecipitation is a precise and powerful assay that can be used to establish the physical interaction of an individual RBP with its target RNAs in vivo. Here, we describe a quantitative method for determining RNA-protein interactions using RNA immunoprecipitation (RNA-IP) assay in mouse embryonic stem cells carrying ectopically expressed mutant constructs. This protocol is reliable and easily adaptable to identify the interactions of endogenous or ectopically expressed RNAs and proteins.