白血病NPM1基因突变快速检测与分型方法的研究  

Methodological Research on Rapid Detection and Genotyping of NPM1 Gene Mutations in Leukemia

作　　者：陈超[1,2] 林珺 龙凯 杨玉春[3] 李志鹏[2] CHEN Chao;LIN Jun;LONG Kai;YANG Yu-Chun;LI Zhi-Peng(Engineering Research Center of Natural Medicine,Ministry of Eduction,Beijing Normal University,Beijing 100875,China;College of Engineering,Beijing Normal University,Zhuhai 519087,Guangdong Province,China;Faculty of Education,Beijing Normal Uni-versity,Beijing 100081,China)

机构地区：[1]北京师范大学珠海校区天然药物教育部工程研究中心,北京100875 [2]北京师范大学珠海分校工程技术学院,广东珠海519087 [3]北京师范大学教育学部中国社会与教育发展研究院,北京100081

出　　处：《中国实验血液学杂志》2021年第2期403-407,共5页Journal of Experimental Hematology

基　　金：北京师范大学教育学部2018年度学科建设综合专项资金资助(2018XKTD003);国家创新训练项目(201813177001)。

摘　　要：目的:建立一种通过荧光探针熔解曲线分析技术对急性髓系白血病(AML)中NPM1突变基因快速检测和分型的方法。方法:设计Nucleoplasmin(NPM1)第12号外显子突变基因mut A、mut B、mut D和野生型的一对共用引物和一条荧光单链探针(分子信标),依据PCR产物与探针的熔解曲线峰值对A、B、D型基因突变进行检测与分型。结果:本方法可对NPM1的A、B、D型基因突变和野生型进行有效分型,灵敏度为1%。通过对62份AML临床样本进行检测,本方法的检出率、分型结果与临床样本测序信息一致。结论:本研究建立的NPM检测方法,PCR体系简单稳定,操作便捷,结果易于判读,可以与测序法互为补充作为AML早期诊断或伴随诊断的NPM1突变快速检测分型方法。Objective:To establish a method for rapid detection and typing of NPM1 mutations in acute myeloid leukemia(AML)by fluorescence melting curve analysis technology.Methods:A pair of primers and a fluorescent single-stranded probe(molecule beacon)were designed for the mutant genes mutA,mutB,mutD in exon 12 of nucleopsin(NPM1)and wild type.With a real-time qPCR,the A,B,and D gene mutations of NPM1 were detected and typed by different-melting curve peak value of the probe through RT-PCR.Results:This method could detected the mutations of A,B,and D in NPM1 effectively with a sensitivity of 1%.Furthermore,62 AML clinical samples were evaluated by the method.In the results,the detection rate and typing of NPM1 mutations were consistent with the sequencing results of clinical samples.Conclusion:There are three features in the method of fluorescence melting curve analysis:stable PCR system,easy to operate,and the easily distinguishable results.The method might meet the demand for rapid typing of NPM1 gene mutation in early diagnosis or concomitant diagnosis of AML.

关 键 词：荧光探针熔解曲线分析 NPM1 检测与分型 

分 类 号：R733.71[医药卫生—肿瘤]