摘要
针对KRAS基因中的2个常见突变G12R和G12S,建立了基于数字聚合酶链反应(PCR)和下一代测序(NGS)技术的特异性检测方法。利用含有目标突变的细胞系配制成不同突变比例的样本,考察了所建立的数字PCR和NGS方法的分析灵敏度和2种方法测量结果的可比性。结果发现对突变比例≥0.83%的样品,NGS和数字PCR的测定结果无显著差异(p>0.05)。突变比例<0.83%的样品,2种方法的测定结果差异显著(p<0.05),而数字PCR方法的测定结果与配制值无显著差异。这表明2种方法在检测高丰度突变时测量结果可比,而数字PCR方法在测定低丰度突变时准确性更高。
Two specific detection methods for KRAS G12R and G12S mutations, based on digital PCR and next generation sequencing （ NGS ）, respectively, were established. Taking the advantage of cell lines containing the target KRAS mutation to prepare a different proportion of G12R and G12S mutations, the analysis sensitivity and comparability of the two proposed methods was investigated. It turned out that no significant difference （p 〉 0. 05 ） was observed between NGS and digital PCR when measuring the sample containing mutation ratio over than 0. 83%. However, the result analyzed by digital PCR was significantly different （p 〈 0. 05 ） from NGS, when measuring the sample containing mutation ratio less than 0. 83%. There was no significant difference between the digital PCR result and the theoretical preparation value. This indicated that the two methods were comparable when analyzing high abundance of KRAS mutation, and digital PCR with higher accuracy is proper applied in the detection of low abundance KRAS mutation. The sensitive of NGS for detection of KRAS mutation is about 1%, whereas digital PCR showed a better sensitivity of 0. 02% when analvzing G12S mutation.
作者
董莲华
王晶
傅博强
段宇航
隋志伟
DONG Lian-Hua, WANG Jing, FU Bo-qiang, DUAN Yu-hang, SUI Zhi-wei(National Institute of Metrology, Beijing 100029, Chin)
出处
《计量学报》
CSCD
北大核心
2018年第3期436-441,共6页
Acta Metrologica Sinica
基金
国家重点研发计划(2016YFC1000301-3)
公益行业专项(AHY1504)
中国计量科学研究院基本业务费(AKY1612)
