Enhanced and controlled chromatin extraction from FFPE tissues and the application to ChIP-seq

Autor: Chad R. Clark, Zhenqing Ye, Jian Zhong, Huihuang Yan, Jeong Heon Lee, Keith D. Robertson, Justin H. Nguyen, Tamas Ordog, Zhiguo Zhang, Samuel W. Lenz, Gianrico Farrugia
Jazyk: angličtina
Rok vydání: 2019
Předmět:
Zdroj: BMC Genomics, Vol 20, Iss 1, Pp 1-11 (2019)
BMC Genomics
ISSN: 1471-2164
DOI: 10.1186/s12864-019-5639-8
Popis: Background Epigenetic dysregulation is involved in the etiology and progression of various human diseases. Formalin-fixed paraffin-embedded (FFPE) samples represent the gold standard for archiving pathology samples, and thus FFPE samples are a major resource of samples in clinical research. However, chromatin-based epigenetic assays in the clinical settings are limited to fresh or frozen samples, and are hampered by low chromatin yield in FFPE samples due to the lack of a reliable and efficient chromatin preparation method. Here, we introduce a new chromatin extraction method from FFPE tissues (Chrom-EX PE) for chromatin-based epigenetic assays. Results During rehydration of FFPE tissues, applying a tissue-level cross-link reversal into the deparaffinized tissue at 65 °C dramatically increased chromatin yield in the soluble fraction. The resulting chromatin is compatible with targeted ChIP-qPCR and genome-wide ChIP-seq approaches. The chromatin prepared by Chrom-EX PE showed a gradual fragmentation pattern with varying incubation temperature. At temperatures below 37 °C, the majority of soluble chromatin is over 1 kb. The soluble chromatin prepared in the range of 45–60 °C showed a typical nucleosomal pattern. And the majority of chromatin prepared at 65 °C is close to mononucleosomal size. These observations indicate that chromatin preparation from FFPE samples can be controlled for downstream chromatin-based epigenetic assays. Conclusions This study provided a new method that achieves efficient extraction of high-quality chromatin suitable for chromatin-based epigenetic assays with less damage on chromatin. This approach may provide a way to circumvent the over-fixed nature of FFPE tissues for future technology development. Electronic supplementary material The online version of this article (10.1186/s12864-019-5639-8) contains supplementary material, which is available to authorized users.
Databáze: OpenAIRE
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