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PCR—Pyrosequencing快速检测药物代谢酶基因CYP2C19多态性方法的建立及应用 被引量：3

Development of a PCR-Pyrosequencing Method for Detecting Rapidly the Genetic Polymorphism of CYP2C19 about Drug Metabolizing Enzymes Genes
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摘要 目的旨在建立一种快速及高通量的CYP2C19药物代谢酶相关基因多态性的检测方法，为个体化治疗提供用药参考。方法抽取96例人外周血DNA，应用带有生物素标记的扩增引物，经PCR扩增并制备焦磷酸测序单链模板，于PyroMarkID焦磷酸测序仪上进行焦磷酸测序，且以Sanger测序法测序结果为对照，观察分析准确性。结果运用焦磷酸测序可以成功检测CYP2C19相关基因型；单个sNP住点检测经与Sanger测序方法比较，符合率为100％。同时，通过设计焦磷酸测序引物，两个PCR产物可合并于一管中进行焦磷酸测序，与单管检测一个位点的焦磷酸测序方法相比，能有效降低检测成本，节省检测时间。结论该方法可准确、高通量、快速检测药物代谢酶相关基因多态性，该方法在个性化医疗上有较大的推广应用价值，也可以将该平台运用于其他疾病相关基因多态性检测。 Objective To develop a new high throughput method for detecting genetic polymorphism of CYP2C19 about drug metabolism enzyme genes rapidly. Methods 96 examples of genomic DNA from human peripheral blood was extracted,and amplified with biotinylated primer to obtain single-stranded templates for pyrosequencing. Then, the single-stranded tem- plates were subjected to Pyrosequencing analysis using PyroMark ID instrument. Simultaneously, Sanger sequencing was also applied to sequence the products as a control to check the reliability of the pyrosequencing result. Results The results displayed that CYP2C19 related genetic polymorphism could be sucessfuly detected using pyrosequencing method, Repeat test and reliability test indicated that the agreement between the pyrosequencing and the Sanger sequencing methods was 100%. Meanwhile,multiple PCR products combined with pyrosequencing detection through design the pyrosequeneing primer; compared with a single-tube sites pyrosequencing methods, it was much more economical and timesaving. Conelution All of these demonstrated that pyrosequencing could accurately and rapidly detect the genetic polymorphism of drug metabolism-related genes with high throughput. It has a great value in personalized medical treatment and could be extended to the other genetic diseases.
作者 施宏 虞闰六 马金飞 任绪义
机构地区 杭州迪安医学检验中心
出处 《现代检验医学杂志》 CAS 2012年第1期66-70,共5页 Journal of Modern Laboratory Medicine
关键词 CYP2C19 焦磷酸测序 CYP2C19 pyrosequencing
分类号 Q786 [生物学—分子生物学]
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参考文献14

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共引文献27

同被引文献23

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现代检验医学杂志
2012年 第1期

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