Cell freezing protocol suitable for ATAC-Seq on motor neurons derived from human induced pluripotent stem cells
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; ... Show more Show less
MetadataShow full item record
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.
DepartmentMassachusetts Institute of Technology. Computational and Systems Biology Program; Massachusetts Institute of Technology. Department of Biological Engineering
Nature Publishing Group
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
Final published version