ATAC‐seq: A Method for Assaying Chromatin Accessibility Genome‐Wide

Jason D. Buenrostro1, Beijing Wu2, Howard Y. Chang1, William J. Greenleaf2

1 Program in Epithelial Biology and the Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, California, 2 Department of Genetics, Stanford University School of Medicine, Stanford, California
Publication Name:  Current Protocols in Molecular Biology
Unit Number:  Unit 21.29
DOI:  10.1002/0471142727.mb2129s109
Online Posting Date:  January, 2015
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Abstract

This unit describes Assay for Transposase‐Accessible Chromatin with high‐throughput sequencing (ATAC‐seq), a method for mapping chromatin accessibility genome‐wide. This method probes DNA accessibility with hyperactive Tn5 transposase, which inserts sequencing adapters into accessible regions of chromatin. Sequencing reads can then be used to infer regions of increased accessibility, as well as to map regions of transcription‐factor binding and nucleosome position. The method is a fast and sensitive alternative to DNase‐seq for assaying chromatin accessibility genome‐wide, or to MNase‐seq for assaying nucleosome positions in accessible regions of the genome. © 2015 by John Wiley & Sons, Inc.

Keywords: ATAC‐seq; transposase; chromatin accessibility

     
 
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Table of Contents

  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
     
 
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Materials

Basic Protocol 1:

  Materials
  • 50,000 cells in a single‐cell suspension
  • Phosphate buffered saline (PBS; appendix 22)
  • Lysis buffer (see recipe)
  • Molecular biology‐grade IGEPAL CA‐630 (Sigma‐Aldrich, cat. no. I8896)
  • TD (2× reaction buffer from Nextera kit; Illumina, cat. no. FC‐121‐1030)
  • TDE1 (Nextera Tn5 Transposase from Nextera kit; Illumina, cat. no. FC‐121‐1030)
  • Nuclease‐free H 2O (available from various molecular biology suppliers)
  • Qiagen MinElute PCR Purification Kit
  • 25 μM PCR Primer 1 [custom‐synthesized by Integrated DNA Technologies (IDT); sequences provided in Buenrostro et al. ( )]
  • 25 μM Barcoded PCR Primer 2 [custom‐synthesized by Integrated DNA Technologies (IDT); sequences provided in Buenrostro et al. ( )]
  • NEBNext High‐Fidelity 2× PCR Master Mix (New England Biolabs, cat. no. M0541)
  • 100× SYBR Green I (Invitrogen, cat. no. S‐7563)
  • 5% TBE polyacrylamide gel (see unit 10.2; optional)
  • 100‐bp DNA ladder (New England Biolabs; optional)
  • Refrigerated centrifuge
  • 0.2‐ml PCR tubes
  • PCR thermal cycler
  • qPCR instrument (Applied Biosystems StepOnePlus Real‐Time PCR System; cat. no. 4376600)
  • Typhoon TRIO Variable Mode Imager (Amersham Biosciences; optional)
  • Bioanalyzer High‐Sensitivity DNA Analysis kit (Agilent; optional)
  • Additional reagents and equipment for counting cells ( appendix 3F), PCR (unit 15.1 and other units in Chapter 15), and polyacrylamide gel electrophoresis (unit 10.2; optional)
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Figures

Videos

Literature Cited

Literature Cited
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