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Single-Molecule Multikilobase-Scale Profiling of Chromatin Accessibility Using m6A-SMAC-Seq and m6A-CpG-GpC-SMAC-Seq

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Chromatin

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2458))

Abstract

A hallmark feature of active cis-regulatory elements (CREs) in eukaryotes is their nucleosomal depletion and, accordingly, higher accessibility to enzymatic treatment. This property has been the basis of a number of sequencing-based assays for genome-wide identification and tracking the activity of CREs across different biological conditions, such as DNAse-seq, ATAC-seq , NOMeseq, and others. However, the fragmentation of DNA inherent to many of these assays and the limited read length of short-read sequencing platforms have so far not allowed the simultaneous measurement of the chromatin accessibility state of CREs located distally from each other. The combination of labeling accessible DNA with DNA modifications and nanopore sequencing has made it possible to develop such assays. Here, we provide a detailed protocol for carrying out the SMAC-seq assay (Single-Molecule long-read Accessible Chromatin mapping sequencing), in its m6A-SMAC-seq and m6A-CpG-GpC-SMAC-seq variants, together with methods for data processing and analysis, and discuss key experimental and analytical considerations for working with SMAC-seq datasets.

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Acknowledgments

The authors thank members of the Greenleaf and Kundaje labs for many helpful discussions. This work was supported by NIH grants UM1HG009436 and P50HG007735 (to W.J.G.). W.J.G. is a Chan Zuckerberg investigator. Z.S. is supported by EMBO Long-Term Fellowship EMBO ALTF 1119-2016 and by Human Frontier Science Program LongTerm Fellowship HFSP LT 000835/2017-L. G.K.M. was supported by the Stanford School of Medicine Dean’s Fellowship.

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Author notes

  1. Georgi K. Marinov and Zohar Shipony contributed equally with all other contributors.

Authors and Affiliations

  1. Department of Genetics, Stanford University, Stanford, CA, USA

    Georgi K. Marinov, Zohar Shipony, Anshul Kundaje & William J. Greenleaf

  2. Department of Computer Science, Stanford University, Stanford, CA, USA

    Anshul Kundaje

  3. Center for Personal Dynamic Regulomes, Stanford University, Stanford, CA, USA

    William J. Greenleaf

  4. Department of Applied Physics, Stanford University, Stanford, CA, USA

    William J. Greenleaf

  5. Chan Zuckerberg Biohub, San Francisco, CA, USA

    William J. Greenleaf

Authors
  1. Georgi K. Marinov
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  2. Zohar Shipony
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  3. Anshul Kundaje
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  4. William J. Greenleaf
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Corresponding authors

Correspondence to Georgi K. Marinov or Zohar Shipony .

Editor information

Editors and Affiliations

  1. Department of Pathology Dunedin School of Medicine, University of Otago, Dunedin, New Zealand

    Julia Horsfield

  2. Cardiology, Division Heart & Lungs, University Medical Center Utrecht, Utrecht, The Netherlands

    Judith Marsman

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© 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Marinov, G.K., Shipony, Z., Kundaje, A., Greenleaf, W.J. (2022). Single-Molecule Multikilobase-Scale Profiling of Chromatin Accessibility Using m6A-SMAC-Seq and m6A-CpG-GpC-SMAC-Seq. In: Horsfield, J., Marsman, J. (eds) Chromatin. Methods in Molecular Biology, vol 2458. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2140-0_15

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  • DOI: https://doi.org/10.1007/978-1-0716-2140-0_15

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2139-4

  • Online ISBN: 978-1-0716-2140-0

  • eBook Packages: Springer Protocols

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