Abstract
DNA accessibility to transcription regulators varies between cells and modulates gene expression patterns. Several “open” chromatin profiling methods that provide valuable insight into the activity of these regulatory regions have been developed. However, their application to clinical samples has been limited despite the discovery that the Analysis of Transposase-Accessible Chromatin followed by sequencing (ATAC-seq) method can be performed using fewer cells than other techniques. Obtaining fresh rather than stored samples and a lack of adequate optimization and quality controls are major barriers to ATAC’s clinical implementation. Here, we describe an optimized ATAC protocol in which we varied nuclear preparation conditions and transposase concentrations and applied rigorous quality control measures before testing fresh, flash frozen, and cryopreserved breast cells and tissue. We obtained high quality data from small cell number. Furthermore, the genomic distribution of sequencing reads, their enrichment at transcription start sites, and transcription factor footprint analyses were similar between cryopreserved and fresh samples. This updated method is applicable to clinical samples, including cells from fine needle aspiration and tissues obtained via core needle biopsy or surgery. Chromatin accessibility analysis using patient samples will greatly expand the range of translational research and personalized medicine by identification of clinically-relevant epigenetic features.
Highlights
The DNA hypersensitivity methods mentioned above require millions of cells, which represents a major obstacle to performing global chromatin landscape analysis using clinical samples, which typically contain less than 100,000 cells
We found that high quality genome-wide open chromatin landscape data that is comparable to that produced using living cells can be generated from a small number of cells, as well as from small tissue samples stored by cryopreservation
Adapting ATAC-seq to small, stored clinical samples will greatly expand the reach of translational research and allow researchers to fully characterize the link between chromatin landscape changes and disease
Summary
The DNA hypersensitivity methods mentioned above require millions of cells, which represents a major obstacle to performing global chromatin landscape analysis using clinical samples, which typically contain less than 100,000 cells. The Analysis of open chromatin sites by Transposase-Accessible Chromatin followed by deep sequencing (ATAC-seq) hypersensitivity method was recently reported to only require small number of fresh cells[11,12,13]. Adapting ATAC-seq to small, stored clinical samples will greatly expand the reach of translational research and allow researchers to fully characterize the link between chromatin landscape changes and disease. This approach may be applicable for personalized medicine therapeutic strategies employed by clinicians treating breast cancer and other diseases
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