Springer Protocols: Abstract: Ribotrap: Targeted Purification of RNA-Specific RNPs from Cell Lysates Through Immunoaffinity Precipitation to Identify Regulatory Proteins and RNAs

Ribotrap: Targeted Purification of RNA-Specific RNPs from Cell Lysates Through Immunoaffinity Precipitation to Identify Regulatory Proteins and RNAs

By: Dale L. Beach^{3 4}, Jack D. Keene⁴

Abstract

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Many elegant methodologies have been devised to explore RNA-protein as well as RNA–RNA interactions. Although the characterization of messages targeted by a specific RNA-binding protein (RBP) has been accelerated by the application of microarray technologies, reliable methods to describe the endogenous assembly of ribonucleoproteins (RNPs) are needed. However, this approach requires the targeted purification of a select mRNA under conditions favorable for the copurification of associated factors including RNA and protein components of the RNP. This chapter describes previous methods used to characterize RNPs in the context of in vitro approaches and presents the Ribotrap methodology, an in vivo protocol for message-specific purification of a target RNP. The method was developed in a yeast model system, yet is amenable to other in vivo cell systems including mammalian cell culture.

Affiliation(s): (3) University of North Carolina, Chapel Hill, USA
(4) Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC, USA

Book Title: Post-Transcriptional Gene Regulation

Series: Methods in Molecular Biology | Volume: 419 | Pub. Date: Dec-21-2007 | Page Range: 69-91 | DOI: 10.1007/978-1-59745-033-1_5

Subject: Genetics/Genomics

Key Words: RNP - ribonomics - RNA-binding protein - post-transcriptional regulation - MS2 - aptamer

References

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