TP53基因是肿瘤的抑制基因,也是癌症中常用的靶向检测基因。R273H和R273C是TP53基因检测中常见的热点突变。为了建立这2个突变的微滴式数字聚合酶链式反应(ddPCR)检测方法,针对突变位点分别设计了2套ddPCR方法,验证了探针的特异性,并对退火温度和引物探针浓度进行了优化。结果显示,所建立的方法具有良好的特异性和重复性,ddPCR法与重量法之间的相关系数为0.9997和0.9996。R273H和R273C的检出限均为0.02%,定量限均为0.05%。这2套方法可用于准确检测TP53-R273H和TP53-R273C基因突变,为临床应用提供支持。
Abstract
TP53 gene is the tumor suppressor gene and commonly used as the targeted detection gene in cancer.R273H and R273C are common hot spot mutations in TP53 gene detection.In order to establish a droplet digital polymerase chain reaction (ddPCR) detection method for these two mutations, two sets of ddPCR methods were designed for the mutation sites, and the specificity of the probe was verified.The annealing temperature and primer probe concentration were optimized.The results showed that the established method had good specificity and repeatability, and the correlation coefficients between ddPCR and weight method were 0.9997 and 0.9996,.The limit of detection of R273H and R273C were both 0.02%, and the limit of quantitation were both 0.05%.The methods can be used to accurately detect TP53-R273H and TP53-R273C gene mutations, providing strong support for clinical application.
关键词
生物计量 /
TP53;基因突变;R273H /
R273C /
核酸检测;数字PCR
Key words
biometrology /
TP53;gene mutation /
R273H /
R273C /
nucleic acid testing;ddPCR
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基金
中国计量科学研究院基本科研业务费重点领域专项(AKYZD202202)
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