扩增阻滞突变系统快速检测H型高血压患者MTHFR C677T基因型的应用研究

Rapid Detection of MTHFR C677T Genotype in H-type Hypertension Patients by Amplification Refractory Mutation System

目的通过扩增阻滞突变系统-聚合酶链反应(ARMS-PCR)检测H型高血压患者5,10-亚甲基四氢叶酸还原酶(MTHFR)C677T基因型,以期建立一种低成本、简单快速的MTHFR C677T基因分型方法。方法基于湖南省H型高血压危险因素研究的大型流行病调查的人群基础,采用单纯随机方法,选取94例H型高血压患者作为研究对象。设计阻滞度递减的系列错配引物,采用ARMS-PCR对94例患者外周血样本的MTHFR C677T基因型进行鉴定,并抽取不同基因型进行测序验证。结果 ARMS-PCR产物电泳后条带清晰,易于判读MTHFR C677T基因型。94例H型高血压患者共检出CC基因型42例、CT基因型24例和TT基因型28例。ARMS-PCR与DNA测序结果一致。结论 ARMS-PCR经阻滞度递减的系列错配引物能有效对MTHFR C677T基因型进行鉴定,此法具有成本低、操作简单、用时短的优点。适用于各类基因的多态性分析。

Objective To develop a cost-effective,rapid and simple method for MTHFR C677 T genotyping by amplification refractory mutation system-polymerase chain reaction（ ARMS-PCR） on H-type hypertension patients.Methods From the population sampled by the large epidemiological investigation based on the research of risk factors for H-type hypertension in Hunan Province,94 H-type hypertension patients were taken as subjects using simple random method. A series of mismatched-primers of refractory reduced were designed,and ARMS-PCR was employed to identify the MTHFR C677 T genotypes of peripheral blood samples of the 94 patients, and different genotypes were abstracted for sequencing validation. Results The bands of ARMS-PCR product agarose gel electrophoretogram were clear sufficiently to identify genotype. Among the 94 H-type hypertension patients 42 were CC genotype,24 were CT genotype and 28 were TT genotype.ARMS-PCR results were in complete accord with DNA sequencing. Conclusion ARMS-PCR is a low-cost,simple and time-saving method for the determination of MTHFR C677 T genotype by mismatched-primers of refractory reduced. The approach can be used to detect other gene polymorphism.