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Global Analysis of Transcription Factor-Binding Sites in Yeast Using ChIP-Seq

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Yeast Genetics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1205))

Abstract

Transcription factors influence gene expression through their ability to bind DNA at specific regulatory elements. Specific DNA-protein interactions can be isolated through the chromatin immunoprecipitation (ChIP) procedure, in which DNA fragments bound by the protein of interest are recovered. ChIP is followed by high-throughput DNA sequencing (Seq) to determine the genomic provenance of ChIP DNA fragments and their relative abundance in the sample. This chapter describes a ChIP-Seq strategy adapted for budding yeast to enable the genome-wide characterization of binding sites of transcription factors (TFs) and other DNA-binding proteins in an efficient and cost-effective way.

Yeast strains with epitope-tagged TFs are most commonly used for ChIP-Seq, along with their matching untagged control strains. The initial step of ChIP involves the cross-linking of DNA and proteins. Next, yeast cells are lysed and sonicated to shear chromatin into smaller fragments. An antibody against an epitope-tagged TF is used to pull down chromatin complexes containing DNA and the TF of interest. DNA is then purified and proteins degraded. Specific barcoded adapters for multiplex DNA sequencing are ligated to ChIP DNA. Short DNA sequence reads (28–36 base pairs) are parsed according to the barcode and aligned against the yeast reference genome, thus generating a nucleotide-resolution map of transcription factor-binding sites and their occupancy.

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Author information

Authors and Affiliations

  1. Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, CT, USA

    Philippe Lefrançois

  2. Department of Biology, West Virginia University, 53 Campus Drive, LSB 5108, Morgantown, WV, 26506, USA

    Jennifer E. G. Gallagher

  3. Department of Genetics, MC: 5120, Stanford University School of Medicine, 300 Pasteur Drive, M-344, Stanford, CA, 94305, USA

    Michael Snyder

Authors
  1. Philippe Lefrançois
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  2. Jennifer E. G. Gallagher
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  3. Michael Snyder
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Corresponding author

Correspondence to Michael Snyder .

Editor information

Editors and Affiliations

  1. Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, Virginia, USA

    Jeffrey S. Smith

  2. Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine, Charlottesville, Virginia, USA

    Daniel J. Burke

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Lefrançois, P., Gallagher, J.E.G., Snyder, M. (2014). Global Analysis of Transcription Factor-Binding Sites in Yeast Using ChIP-Seq. In: Smith, J., Burke, D. (eds) Yeast Genetics. Methods in Molecular Biology, vol 1205. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1363-3_15

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  • DOI: https://doi.org/10.1007/978-1-4939-1363-3_15

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1362-6

  • Online ISBN: 978-1-4939-1363-3

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