Improved results of LINE-1 methylation analysis in formalin-fixed, paraffin-embedded tissues with the application of a heating step during the DNA extraction process
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- 作者：Xianyu Wen ; Seorin Jeong ; Younghoon Kim ; Jeong Mo Bae ; Nam Yun Cho…
- 关键词：Archival tissue ; CpG island methylator phenotype ; DNA methylation ; Formalin ; Heat treatment ; LINE ; 1
- 刊名：Clinical Epigenetics
- 出版年：2017
- 出版时间：December 2017
- 年：2017
- 卷：9
- 期：1
- 全文大小：941KB
- 刊物主题：Human Genetics; Gene Function;
- 出版者：BioMed Central
- ISSN：1868-7083
- 卷排序：9
文摘
BackgroundFormalin-fixed, paraffin-embedded (FFPE) tissues are important resources for profiling DNA methylation changes and for studying a variety of diseases. However, formalin fixation introduces inter-strand crosslinking, which might cause incomplete bisulfite conversion of unmethylated cytosines, which might lead to falsely elevated measurements of methylation levels in pyrosequencing assays. Long interspersed nucleotide element-1 (LINE-1) is a major constituent of repetitive transposable DNA elements, and its methylation is referred to correlates with global DNA methylation. To identify whether formalin fixation might impact the measured values of methylation in LINE-1 repetitive elements and whether prolonged heat-induced denaturation of DNA might reduce the artificial increases in measured values caused by formalin fixation, we analyzed paired fresh-frozen (FF) and FFPE xenograft tissue samples for their methylation levels in LINE-1 using a pyrosequencing assay. To further confirm the effect of a heating step in the measurement of LINE-1 or single gene methylation levels, we analyzed FFPE tissue samples of gastric cancer and colorectal cancer for their methylation status in LINE-1 and eight single genes, respectively.