Cell freezing protocol optimized for ATAC-Seq on motor neurons derived from human induced pluripotent stem cells

BioRxiv : the Preprint Server for Biology
Pamela MilaniErnest Fraenkel

Abstract

In recent years, the assay for transposase-accessible chromatin using sequencing (ATAC-Seq) has become a fundamental tool of epigenomic research. However, it has proven difficult to perform this technique on frozen samples because freezing cells before extracting nuclei impairs nuclear integrity and alters chromatin structure. We describe a protocol for freezing cells that is compatible with ATAC-Seq, producing results that compare well with those generated from fresh cells. We found that while flash-frozen samples are not suitable for ATAC-Seq, the assay is successful with slow-cooled cryopreserved samples. Using this method, we were able to isolate high quality, intact nuclei, and we verified that epigenetic results from fresh and cryopreserved samples agree quantitatively. We developed our protocol on a disease-relevant cell type, namely motor neurons differentiated from induced pluripotent stem cells from a patient affected by spinal muscular atrophy.

Related Concepts

Research
Transposase
Pluripotent Stem Cells
Spinal Muscular Atrophy
XCL1
Human Induced Pluripotent Stem Cells
Nucleic Acid Sequencing
Motor Neurons
Cell Nucleus
Cryopreservation

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