An RNA-immunoprecipitation protocol to identify RNAs associated with RNA-binding proteins in cytoplasmic and nuclear Drosophila head fractions

Abstract

Summary: RNA-binding proteins (RBPs) are multifunctional proteins that shuttle between the nucleus and the cytoplasm where they assemble with target RNAs to form multi-molecular complexes. Here, we describe a protocol to selectively identify RNAs associated with RBPs of interest in the cytoplasmic and nuclear compartments of adult Drosophila brain cells. Cytoplasmic and nuclear fractions are differentially collected and used for immunoprecipitation-based purification of GFP-tagged RBPs. This protocol can be applied to samples expressing ectopic or endogenous tagged RBPs. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell separation/fractionation
• Gene Expression
• Model Organisms
• Molecular Biology
• Protein Biochemistry