Cell freezing protocol suitable for ATAC-Seq on motor neurons derived from human induced pluripotent stem cells
Name
Cell freezing.pdf
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1.14 MB
Format
Adobe PDF
Checksum (MD5)
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Author(s)
Shelley, Brandon C.
Mandefro, Berhan
Sareen, Dhruv
Svendsen, Clive N.
Milani, Pamela
Escalante, Renan A.
Patel-Murray, Natasha Leanna
Xin, Xiaofeng
Adam, Miriam
Fraenkel, Ernest
Date Issued
May 2016
Journal
Scientific Reports
Publisher
Nature Publishing Group
Citation
Milani, Pamela et al. “Cell Freezing Protocol Suitable for ATAC-Seq on Motor Neurons Derived from Human Induced Pluripotent Stem Cells.” Scientific Reports 6.1 (2016): n. pag. © 2017 Macmillan Publishers Limited
Version
Final published version
Abstract
In recent years, the assay for transposase-accessible chromatin using sequencing (ATAC-Seq) has become a fundamental tool of epigenomic research. However, it is difficult to perform this technique on frozen samples because freezing cells before extracting nuclei can impair nuclear integrity and alter chromatin structure, especially in fragile cells such as neurons. Our aim was to develop a protocol for freezing neuronal cells that is compatible with ATAC-Seq; we focused on a disease-relevant cell type, namely motor neurons differentiated from induced pluripotent stem cells (iMNs) from a patient affected by spinal muscular atrophy. We found that while flash-frozen iMNs are not suitable for ATAC-Seq, the assay is successful with slow-cooled cryopreserved cells. Using this method, we were able to isolate high quality, intact nuclei, and we verified that epigenetic results from fresh and cryopreserved iMNs quantitatively agree.
MIT Department
Massachusetts Institute of Technology. Computational and Systems Biology Program
Massachusetts Institute of Technology. Department of Biological Engineering
Terms of Use
Creative Commons Attribution 4.0 International License
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DOI of Published Version
https://doi.org/10.1038/srep25474
