Islam Farhadul, Tang Johnny C, Gopalan Vinod, Lam Alfred K
Cancer Molecular Pathology of School of Medicine, Griffith University, Gold Coast, Australia.
Department of Biochemistry and Molecular Biology, University of Rajshahi, Rajshahi, Bangladesh.
Methods Mol Biol. 2018;1756:247-256. doi: 10.1007/978-1-4939-7734-5_21.
The aberrant DNA methylation has been noted to occur at promoter of tumor suppressor, cell adhesion, DNA repair, and other growth regulating genes during the progression of nonneoplastic esophageal mucosa to Barrett esophagus to esophageal adenocarcinoma. Methylation-mediated silencing of individual gene or concurrent loss of a number of genes plays crucial roles in dysplasia-metaplasia-neoplasia sequence of esophageal adenocarcinoma. In addition, promoter methylation of genes had shown significant prognostic potential in patients with esophageal adenocarcinoma. Thus, determination of methylation status of genes of interest can be used as a molecular marker for risk stratification and/or better prognosis of patients with esophageal adenocarcinoma. There are a number of methods including bead array, PCR and sequencing, pyrosequencing, methylation-specific PCR, and PCR with high-resolution melt curve available to determine the methylation status of particular gene of interest. Herein, we describe the polymerase chain reaction followed by sequencing-based protocol for identifying DNA methylation status in esophageal adenocarcinoma.
在非肿瘤性食管黏膜向巴雷特食管再到食管腺癌的进展过程中，已发现异常DNA甲基化发生在肿瘤抑制基因、细胞黏附基因、DNA修复基因以及其他生长调节基因的启动子区域。单个基因的甲基化介导沉默或多个基因的同时缺失在食管腺癌的发育异常-化生-肿瘤形成序列中起着关键作用。此外，基因的启动子甲基化在食管腺癌患者中显示出显著的预后潜力。因此，确定感兴趣基因的甲基化状态可作为食管腺癌患者风险分层和/或更好预后的分子标志物。有多种方法可用于确定特定感兴趣基因的甲基化状态，包括珠阵列、PCR和测序、焦磷酸测序、甲基化特异性PCR以及高分辨率熔解曲线PCR。在此，我们描述了基于测序的聚合酶链反应方案，用于鉴定食管腺癌中的DNA甲基化状态。
