Chapter title |
Analysis of HIV-1 Gag-RNA Interactions in Cells and Virions by CLIP-seq.
|
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Chapter number | 8 |
Book title |
HIV Protocols
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Published in |
Methods in molecular biology, January 2016
|
DOI | 10.1007/978-1-4939-3046-3_8 |
Pubmed ID | |
Book ISBNs |
978-1-4939-3045-6, 978-1-4939-3046-3
|
Authors |
Kutluay, Sebla B, Bieniasz, Paul D, Sebla B. Kutluay, Paul D. Bieniasz, Kutluay, Sebla B., Bieniasz, Paul D. |
Abstract |
Next-generation sequencing-based methodologies have revolutionized the analysis of protein-nucleic acid complexes; yet these novel approaches have rarely been applied in virology. Because it has an RNA genome, RNA-protein interactions play critical roles in human immunodeficiency virus type 1 (HIV-1) replication. In many cases, the binding sites of proteins on HIV-1 RNA molecules in physiologically relevant settings are not known. Cross-linking-immunoprecipitation sequencing (CLIP-seq) methodologies, which combine immunoprecipitation of covalently crosslinked protein-RNA complexes with high-throughput sequencing, is a powerful technique that can be applied to such questions as it provides a global account of RNA sequences bound by a RNA-binding protein of interest in physiological settings at near-nucleotide resolution. Here, we describe the application of the CLIP-seq methodology to identify the RNA molecules that are bound by the HIV-1 Gag protein in cells and in virions. This protocol can easily be applied to other viral and cellular RNA-binding proteins that influence HIV-1 replication. |
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