RNA Immunoprecipitation Assay to Determine the Specificity of SRSF3 Binding to Nanog mRNA

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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 immuno precipitation is a precise and powerful assay that can be used to establish the physical interaction of an individual RBP with its target RNAs invivo. 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.
Original languageEnglish
Article numbere3071
Number of pages9
JournalBio-protocol
Volume8
Issue number21
DOIs
Publication statusPublished - 5 Nov 2018

Keywords

  • RNA immunoprecipitation
  • RNA binding protein
  • SR protein
  • RNA-protein interaction
  • antibody
  • quantitative PCR

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