应用液滴数字PCR确认MSH2基因中的一个新的无-第三军医大学学报.docx

应用液滴数字PCR确认MSH2基因中的一个新的无义基因突变：定性及定量分析潘琦1，丁克越1(1 400010重庆，重庆医科大学附属第二医院感染科、重庆医科大学病毒性肝炎研究所)[摘要] 目的 探讨液滴数字PCR（droplet digital PCR，ddPCR）验证一例原发性肝细胞肝癌患者癌基因组MSH2基因中发生的一个新的无义突变的分析方法。方法 我们对已经通过全外显子组测序鉴定到的一个新的MSH2的体细胞突变进行ddPCR验证；分别对该患者肝细胞肝癌组织和癌旁硬化组织的基因组DNA分析。结果 ddPCR定性验证了该无义突变 (r g.chr2CT, c.28 CT, p.Q10*) 只发生在肝癌组织并非在癌旁硬化组织中，定量研究发现该无义突变在肝癌组织中的等位基因分数为0.39，与外显子测序的结果类似。结论 ddPCR是一种快速和准确的基因突变分析的新方法，灵敏度和准确性高，在基因突变的分析中具有广泛应用。[关键词] 微滴数字PCR（ddPCR）；肝细胞肝癌；MSH2基因；无义突变（nonsense mutation）[中图法分类号] X703.1；R394.3；R733.71 Droplet digital PCR as an accurate method for validating a novel nonsense somatic mutation in MSH2: qualitatively and quantitativelyPan Qi1, Ding Keyue1（1 Institute of Viral Hepatitis, Key Laboratory of Molecular Biology for Infectious Diseases, Ministry of Education of China, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, China）[Abstract] Objective: To validate a novel nonsense somatic mutation in MSH2 of a patient with primary hepatocellular carcinoma using droplet digital PCR. Method: ddPCR was used to validate a novel nonsense mutation in MSH2 identified by whole exome sequencing. We performed qualitative and quantitative analysis in the liver cancer tissues and adjacent non-cancerous tissues. Result: ddPCR validated that the nonsense mutation (r g. CT, p.Q10*) in MSH2 occurred in the liver cancer tissues rather than the adjacent non-cancerous tissues. The allelic fraction of the nonsense mutation obtained from ddPCR was similar with that estimated from the whole-exome sequencing. Conclusion: ddPCR is a new, rapid and accurate method for mutation analysis with a high sensitivity and accuracy.[Keywords] Droplet digital PCR；Hepatocellular carcinoma；MSH2；Nonsense mutationSupported by the Recruitment Program of Global Youth Experts in China (KD).Corresponding author: Keyue Ding, Tel: +86-23 E-mail: ding.keyue@cqmu.edu.cn数字PCR (digital PCR) 是一种新型的DNA分子计数技术，可直接计算样品中DNA靶分子和标准分子的绝对数值{Vogelstein:1999ve}。随着