State Key Laboratory of Transducer Technology, Shanghai Institute of Microsystem and Information Technology, Chinese Academy of Sciences, Shanghai 200050, China; University of Chinese Academy of Sciences, Beijing 100039, China.
State Key Laboratory of Transducer Technology, Shanghai Institute of Microsystem and Information Technology, Chinese Academy of Sciences, Shanghai 200050, China.
Biosens Bioelectron. 2017 Oct 15;96:339-344. doi: 10.1016/j.bios.2017.05.021. Epub 2017 May 12.
Hypermethylation of CpG islands in the promoter region of many tumor suppressor genes downregulates their expression and in a result promotes tumorigenesis. Therefore, detection of DNA methylation status is a convenient diagnostic tool for cancer detection. Here, we reported a novel method for the integrative detection of methylation by the microfluidic chip-based digital PCR. This method relies on methylation-sensitive restriction enzyme HpaII, which cleaves the unmethylated DNA strands while keeping the methylated ones intact. After HpaII treatment, the DNA methylation level is determined quantitatively by the microfluidic chip-based digital PCR with the lower limit of detection equal to 0.52%. To validate the applicability of this method, promoter methylation of two tumor suppressor genes (PCDHGB6 and HOXA9) was tested in 10 samples of early stage lung adenocarcinoma and their adjacent non-tumorous tissues. The consistency was observed in the analysis of these samples using our method and a conventional bisulfite pyrosequencing. Combining high sensitivity and low cost, the microfluidic chip-based digital PCR method might provide a promising alternative for the detection of DNA methylation and early diagnosis of epigenetics-related diseases.
CpG 岛在许多肿瘤抑制基因启动子区域的超甲基化下调其表达，从而促进肿瘤发生。因此，检测 DNA 甲基化状态是癌症检测的一种便捷的诊断工具。在这里，我们报道了一种基于微流控芯片的数字 PCR 进行甲基化综合检测的新方法。该方法依赖于甲基化敏感的限制性内切酶 HpaII，它可以切割未甲基化的 DNA 链，而保持甲基化的 DNA 链完整。经过 HpaII 处理后，通过微流控芯片数字 PCR 定量测定 DNA 甲基化水平，检测下限可达 0.52%。为了验证该方法的适用性，我们在 10 例早期肺腺癌及其相邻非肿瘤组织的样本中检测了两个肿瘤抑制基因（PCDHGB6 和 HOXA9）的启动子甲基化。使用我们的方法和常规亚硫酸氢盐焦磷酸测序对这些样本进行分析，结果具有一致性。该基于微流控芯片的数字 PCR 方法结合了高灵敏度和低成本的优点，可能为 DNA 甲基化的检测和与表观遗传学相关疾病的早期诊断提供一种有前途的替代方法。
