Posted in Research

A Pipeline for Analyzing eCLIP and iCLIP Data with Htseq-clip and DEWSeq

 October 26, 2021



doi: 10.1007/978-1-0716-1851-6_10.

Affiliations

Item in Clipboard

Sudeep Sahadevan et al.


Methods Mol Biol.


2022.

Abstract

Individual-nucleotide crosslinking and immunoprecipitation (iCLIP) sequencing and its derivative enhanced CLIP (eCLIP) sequencing are methods for the transcriptome-wide detection of binding sites of RNA-binding proteins (RBPs). This chapter provides a stepwise tutorial for analyzing iCLIP and eCLIP data with replicates and size-matched input (SMI) controls after read alignment using our open-source tools htseq-clip and DEWSeq. This includes the preparation of gene annotation, extraction, and preprocessing of truncation sites and the detection of significantly enriched binding sites using a sliding window based approach suitable for different binding modes of RBPs.


Keywords:

Enhanced individual-nucleotide crosslinking and Immunoprecipitation; Next-generation sequencing; RBP binding-site detection; RNA-binding proteins; eCLIP; iCLIP; seCLIP.

References

    1. Gebauer F, Schwarzl T, Valcárcel J, Hentze MW (2020) RNA-binding proteins in human genetic disease. Nat Rev Genet 22:185–198

      –

      DOI

    1. König J, Zarnack K, Rot G et al (2010) ICLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol 17:909–915

      –

      DOI

    1. Van Nostrand EL, Pratt GA, Shishkin AA et al (2016) Robust transcriptome-wide discovery of RNA-binding protein binding sites with enhanced CLIP (eCLIP). Nat Methods 13:508–514

      –

      DOI

    1. Hafner M, Landthaler M, Burger L et al (2010) Transcriptome-wide identification of RNA-binding protein and MicroRNA target sites by PAR-CLIP. Cell 141:129–141

      –

      DOI

    1. Licatalosi DD, Mele A, Fak JJ et al (2008) HITS-CLIP yields genome-wide insights into brain alternative RNA processing. Nature 456:464–469

      –

      DOI

Read more here: Source link