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Polycomb Group Proteins pp 73–86Cite as

RIP: RNA Immunoprecipitation

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1480))

Abstract

The relevance of RNA-protein interactions in modulating mRNA and noncoding RNA function is increasingly appreciated and several methods have been recently developed to map them. The RNA immunoprecipitation (RIP) is a powerful method to study the physical association between individual proteins and RNA molecules in vivo. The basic principles of RIP are very similar to those of chromatin immunoprecipitation (ChIP), a largely used tool in the epigenetic field, but with some important caveats. The approach is based on the use of a specific antibody raised against the protein of interest to pull down the RNA-binding protein (RBP) and target-RNA complexes. Any RNA that is associated with this protein complex will also be isolated and can be further analyzed by polymerase chain reaction-based methods, hybridization, or sequencing.

Several variants of this technique exist and can be divided into two main classes: native and cross-linked RNA immunoprecipitation. The native RIP allows to reveal the identity of RNAs directly bound by the protein and their abundance in the immunoprecipitated sample, while cross-linked RIP leads to precisely map the direct and indirect binding site of the RBP of interest to the RNA molecule.

In this chapter both the protocols applied to mammalian cells are described taking into account the caveats and considerations required for designing, performing, and interpreting the results of these experiments.

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Authors and Affiliations

  1. Institute of Genetics and Biophysics “ABT”- CNR, via Pietro Castellino 111, 80131, Naples, Italy

    Miriam Gagliardi & Maria R. Matarazzo

Authors
  1. Miriam Gagliardi
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  2. Maria R. Matarazzo
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Corresponding author

Correspondence to Maria R. Matarazzo .

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Editors and Affiliations

  1. IBCN-CNR, Institute of Cell Biology and Neurobiology and Istituto Nazionale di Genetica Molecolare “Romeo ed Enrica Invernizzi”, Rome, Italy

    Chiara Lanzuolo

  2. Istituto Nazionale di Genetica Molecolare “Romeo ed Enrica Invernizzi”, Milan, Italy

    Beatrice Bodega

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Gagliardi, M., Matarazzo, M.R. (2016). RIP: RNA Immunoprecipitation. In: Lanzuolo, C., Bodega, B. (eds) Polycomb Group Proteins. Methods in Molecular Biology, vol 1480. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6380-5_7

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  • DOI: https://doi.org/10.1007/978-1-4939-6380-5_7

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6378-2

  • Online ISBN: 978-1-4939-6380-5

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