Robust transcriptome-wide discovery of RNA-binding protein binding sites with enhanced CLIP (eCLIP).

TitleRobust transcriptome-wide discovery of RNA-binding protein binding sites with enhanced CLIP (eCLIP).
Publication TypeJournal Article
Year of Publication2016
AuthorsVan Nostrand EL, Pratt GA, Shishkin AA, Gelboin-Burkhart C, Fang MY, Sundararaman B, Blue SM, Nguyen TB, Surka C, Elkins K, Stanton R, Rigo F, Guttman M, Yeo GW
JournalNat Methods
Volume13
Issue6
Pagination508-14
Date Published2016 Jun
ISSN1548-7105
Abstract

As RNA-binding proteins (RBPs) play essential roles in cellular physiology by interacting with target RNA molecules, binding site identification by UV crosslinking and immunoprecipitation (CLIP) of ribonucleoprotein complexes is critical to understanding RBP function. However, current CLIP protocols are technically demanding and yield low-complexity libraries with high experimental failure rates. We have developed an enhanced CLIP (eCLIP) protocol that decreases requisite amplification by ∼1,000-fold, decreasing discarded PCR duplicate reads by ∼60% while maintaining single-nucleotide binding resolution. By simplifying the generation of paired IgG and size-matched input controls, eCLIP improves specificity in the discovery of authentic binding sites. We generated 102 eCLIP experiments for 73 diverse RBPs in HepG2 and K562 cells (available at https://www.encodeproject.org), demonstrating that eCLIP enables large-scale and robust profiling, with amplification and sample requirements similar to those of ChIP-seq. eCLIP enables integrative analysis of diverse RBPs to reveal factor-specific profiles, common artifacts for CLIP and RNA-centric perspectives on RBP activity.

DOI10.1038/nmeth.3810
PubMed URLhttp://www.ncbi.nlm.nih.gov/pubmed/27018577?dopt=Abstract
Alternate JournalNat. Methods
PubMed ID27018577
PubMed Central IDPMC4887338
Track(s): 
Bioinformatics and Systems Biology