一种用于检测葡萄糖-6-磷酸脱氢酶（G6PD）基因突变的多重方法。
A multiplex method for detection of glucose-6-phosphate dehydrogenase (G6PD) gene mutations.
作者信息
Zhang L, Yang Y, Liu R, Li Q, Yang F, Ma L, Liu H, Chen X, Yang Z, Cui L, He Y
机构信息
Department of Cell Biology and Medical Genetics, Kunming Medical University, Kunming, Yunnan Province, China.
The First Affiliated Hospital, Kunming Medical University, Kunming, Yunnan Province, China.
出版信息
Int J Lab Hematol. 2015 Dec;37(6):739-45. doi: 10.1111/ijlh.12405. Epub 2015 Jul 20.
INTRODUCTION
Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common human enzyme defect caused by G6PD gene mutations. This study aimed to develop a cost-effective, multiplex, genotyping method for detecting common mutations in the G6PD gene.
METHODS
We used a SNaPshot approach to genotype multiple G6PD mutations that are common to human populations in South-East Asia. This assay is based on multiplex PCR coupled with primer extension reactions. Different G6PD gene mutations were determined by peak retention time and colors of the primer extension products.
RESULTS
We designed PCR primers for multiplex amplification of the G6PD gene fragments and for primer extension reactions to genotype 11 G6PD mutations. DNA samples from a total of 120 unrelated G6PD-deficient individuals from the China-Myanmar border area were used to establish and validate this method. Direct sequencing of the PCR products demonstrated 100% concordance between the SNaPshot and the sequencing results.
CONCLUSION
The SNaPshot method offers a specific and sensitive alternative for simultaneously interrogating multiple G6PD mutations.
引言
葡萄糖-6-磷酸脱氢酶（G6PD）缺乏症是由G6PD基因突变引起的最常见的人类酶缺陷。本研究旨在开发一种经济高效的多重基因分型方法，用于检测G6PD基因中的常见突变。
方法
我们采用SNaPshot方法对东南亚人群中常见的多种G6PD突变进行基因分型。该检测基于多重PCR结合引物延伸反应。通过引物延伸产物的峰保留时间和颜色确定不同的G6PD基因突变。
结果
我们设计了用于G6PD基因片段多重扩增和引物延伸反应的PCR引物，以对11种G6PD突变进行基因分型。使用来自中国-缅甸边境地区总共120名无关的G6PD缺乏个体的DNA样本建立并验证了该方法。PCR产物的直接测序表明SNaPshot与测序结果之间的一致性为100%。
结论
SNaPshot方法为同时检测多种G6PD突变提供了一种特异且灵敏的替代方法。
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