Method

Quantifying ChIP-seq data: a spiking method providing an internal reference for sample-to-sample normalization

    • 1Center for Integrative Genomics, Faculty of Biology and Medicine, University of Lausanne, 1015 Lausanne, Switzerland;
    • 2Bioinformatics Core Facility, SIB Swiss Institute of Bioinformatics, 1015 Lausanne, Switzerland;
    • 3Swiss Institute of Bioinformatics, 1015 Lausanne, Switzerland;
    • 4Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York 10461, USA;
    • 5Department of Oncology and the Ludwig Center for Cancer Research, Faculty of Biology and Medicine, University of Lausanne, 1011 Lausanne, Switzerland
    • 1Center for Integrative Genomics, Faculty of Biology and Medicine, University of Lausanne, Lausanne 1015, Switzerland
    • 2Swiss Institute of Bioinformatics, Lausanne, Switzerland
    • 3Bioinformatics Core Facility, Swiss Institute of Bioinformatics, Lausanne, Switzerland
    • 4Department of Oncology and Ludwig Center for Cancer Research, Faculty of Biology and Medicine, University of Lausanne, Switzerland
    • 5Interfaculty Institute of Bioengineering, School of Life Sciences, Ecole polytechnique Fédérale de Lausanne, Switzerland
    • 6Vital IT, Swiss Institute of Bioinformatics, Lausanne, Switzerland
    • 7Bioinformatics and Biostatistics Core Facility, School of Life Sciences, Ecole polytechnique Fédérale de Lausanne, Switzerland
    • 8Department of Molecular Biology, Faculty of Sciences, University of Geneva, Switzerland
    • 6 These authors contributed equally to this work.
    • 7 A complete list of consortium authors appears at the end of this article.
    • 8 Present address: Biocartis SA, EPFL Innovation Square, Lausanne, Switzerland
    • 9 Corresponding authors E-mail [email protected] E-mail [email protected]
Published April 7, 2014. https://doi.org/10.1101/gr.168260.113
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cover of Genome Research Vol 36 Issue 7
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Abstract

Chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) experiments are widely used to determine, within entire genomes, the occupancy sites of any protein of interest, including, for example, transcription factors, RNA polymerases, or histones with or without various modifications. In addition to allowing the determination of occupancy sites within one cell type and under one condition, this method allows, in principle, the establishment and comparison of occupancy maps in various cell types, tissues, and conditions. Such comparisons require, however, that samples be normalized. Widely used normalization methods that include a quantile normalization step perform well when factor occupancy varies at a subset of sites, but may miss uniform genome-wide increases or decreases in site occupancy. We describe a spike adjustment procedure (SAP) that, unlike commonly used normalization methods intervening at the analysis stage, entails an experimental step prior to immunoprecipitation. A constant, low amount from a single batch of chromatin of a foreign genome is added to the experimental chromatin. This “spike” chromatin then serves as an internal control to which the experimental signals can be adjusted. We show that the method improves similarity between replicates and reveals biological differences including global and largely uniform changes.

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