高通量测定技术检测结核分枝杆菌耐利福平的研究

被引：5

作者：

赵锦荣

白玉杰

王琰

张庆华

罗明

闫小君

机构：

[1] 第四军医大学药学系基因诊断技术应用研究所,第四军医大学药学系基因诊断技术应用研究所,第四军医大学药学系基因诊断技术应用研究所,第四军医大学药学系基因诊断技术应用研究所,西安市结核病医院检验科,第四军医大学药学系基因诊断技术应用研究所西安,西安,西安,西安,西安

来源：

中华结核和呼吸杂志 | 2005年 / 05期

关键词：

分枝杆菌,结核; 利福平; 抗药性,微生物; DNA突变分析;

D O I：

暂无

中图分类号：

R446.5 [微生物学检验];

学科分类号：

摘要：

目的基于焦磷酸测序(pyrosequencing)技术,建立耐利福平结核分枝杆菌快速检测方法,并探讨其临床应用价值。方法设计一对聚合酶链反应(PCR)引物,其中一引物经生物素化修饰,PCR扩增含耐药决定区一297bp长的rpoB基因片段,借用链亲和素包被磁珠纯化单链PCR产物。设计2个正向测序引物,运用pyrosequencing技术对耐药决定区进行序列测定。通过对一系列10倍稀释的结核分枝杆菌H37Rv标准株DNA进行分析,评价所建立方法的检测敏感性。通过对已知耐药表型的结核分枝杆菌临床分离株进行分析,评价所建立方法的检测特异性。结果PCR扩增后,可在2h内得到耐药决定区序列。所建立方法的检测灵敏度为50fgDNA/反应。在所分析的利福平敏感株中均未检测到耐药决定区突变,而在耐利福平菌株中均检测到耐药决定区突变。结论所建立的方法具有自动化程度高和结果准确等特点,适用于对耐利福平结核分枝杆菌进行快速高通量检测。

引用

页码：297 / 300

页数：4

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