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目的建立表皮生长因子受体(Epidermal Growth Factor Receptor,EGFR)21号外显子基因突变的焦磷酸测序方法。方法用纯野生和纯突变的EGFR21质粒DNA按比例配制7份不同突变比例的DNA样本和1份正常人血液样本,对焦磷酸测序技术检测EGFR21基因突变方法进行验证,制备175例非小细胞肺癌标本g DNA,应用Qigene Pyro Mark Q24焦磷酸测序仪进行EGFR21基因的焦磷酸测序。结果建立了可同时检测EGFR基因21号外显子L858R和L861Q突变的焦磷酸测序方法,175例非小细胞肺癌标本中共检测出EGFR21突变型样本29例,总突变率为16.57%(29/175),其中L858R突变占14.29%(25/175),L861Q突变占2.28%(4/175)。结论 EGFR21基因突变的焦磷酸测序新方法具有快速、简便、灵敏度和准确度高的优点,特别适合于科研和临床批量检测的需要。
Objective To establish a pyrosequencing method for detecting the mutation of exon 21 gene of epidermal growth factor receptor (EGFR). Methods Seven DNA samples with different mutation ratios and one normal human blood sample were prepared proportionally from wild-type and pure mutant EGFR21 plasmid DNA. The mutation of EGFR21 gene was detected by pyrosequencing technology and 175 non-small cell lung cancer samples g DNA, Pyrosequencing of the EGFR21 gene was performed using the Qigene Pyro Mark Q24 Pyrosequencing System. Results A pyrosequencing method was developed for the simultaneous detection of mutations of L858R and L861Q in exon 21 of EGFR gene. Twenty-nine of 175 non-small cell lung cancer samples were detected with EGFR21 mutation, with a total mutation rate of 16.57% (29/175) , Of which L858R mutation accounted for 14.29% (25/175) and L861Q mutation accounted for 2.28% (4/175). Conclusion The new method of pyrosequencing with EGFR21 mutation is rapid, simple, sensitive and accurate. It is especially suitable for the needs of scientific research and clinical testing.