Abstract
During the last decade a wide range of single-cell and single-nucleus next-generation sequencing techniques have been developed, which revolutionized detection of rare cell populations, enabling creation of comprehensive cell atlases of complex organs and tissues. State-of-the-art methods do not only allow classical transcriptomics of individual cells but also comprise a number of epigenetic approaches, including assessment of chromatin accessibility by single-nucleus Assay for Transposase Accessible Chromatin ATAC-seq (snATAC-seq). The snATAC-seq assay detects “open chromatin,” a term for low nucleosome occupancy of genomic regions, which is a prerequisite for effective transcription factor binding. Information about open chromatin at the single-nucleus level helps to recognize epigenetic changes, sometimes before transcription of respective genes occurs. snATAC-seq detects cellular heterogeneity in otherwise still transcriptionally and/or morphologically homogeneous cell populations. Chromatin accessibility assays may be used to detect epigenetic changes in cardiac lineages during heart development, chromatin landscape changes during aging, and epigenetic alterations in heart diseases. Here, we provide an optimized protocol for snATAC-seq of murine hearts. We describe isolation of single nuclei from snap-frozen hearts, provide hints for preparation of libraries suitable for snATAC-seq next-generation sequencing (NGS) using the Chromium 10× platform, and give general recommendations for downstream analysis using conventional bioinformatic pipelines and packages. The protocol should serve as a beginner’s guide to generate high-quality snATAC-seq datasets and to perform chromatin accessibility analysis of individual heart-derived cell nuclei.
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Acknowledgments
This work was supported by the Excellence Initiative “Cardiopulmonary Institute” (CPI), the DFG collaborative research center SFB1213, Transregional Collaborative Research Centre 267, the DFG Transregional Collaborative Research Centre 81, and the DFG Clinical Research Unit FKO 309.
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Yekelchyk, M., Li, X., Guenther, S., Braun, T. (2024). Single-Nucleus ATAC-seq for Mapping Chromatin Accessibility in Individual Cells of Murine Hearts. In: Gužvić, M. (eds) Single Cell Analysis. Methods in Molecular Biology, vol 2752. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3621-3_16
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DOI: https://doi.org/10.1007/978-1-0716-3621-3_16
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