High-throughput SNP genotyping based on solid-phase PCR on magnetic nanoparticles with dual-color hybridization
High-throughput SNP genotyping based on solid-phase PCR on magnetic nanoparticles with dual-color hybridization
复制标题
基于磁性纳米颗粒固相 PCR 双色杂交的高通量 SNP 基因分型
DOI：
10.1016/j.jbiotec.2007.06.023
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发表时间：
2007-09-15
影响因子：
4.1
通讯作者：
He, Nongyue
中科院分区：
文献类型：
--
作者：
Liu, Hongna;Liab, Song;He, Nongyue
单核苷酸多态性（SNPs）是DNA序列中的一个碱基变异，在试图找到遗传性疾病的基因时通常会有所帮助。在本文中，基于微阵列的分型单核苷酸多态性（SNPs）的方法，使用固相聚合酶链反应（PCR）上的磁性纳米粒子（MNP）。用链霉亲和素包被的磁性纳米颗粒（SA-MNP）捕获一条带有生物素标记的引物，并在96孔板中在SA-MNP表面直接扩增PCR产物。通过与一对双色探针杂交来询问样品以确定SNP，然后通过扫描用变性荧光探针打印的微阵列来同时鉴定每个样品的基因型。应用该方法检测了126例正常人亚甲基四氢叶酸还原酶（MTHFR）基因C677 T多态性。结果表明，三种不同的基因型在芯片上区分三个不同的荧光模式。该方法不需要纯化和还原步骤，所有反应都可以在同一个容器中进行，是一种简单、省力的SNP分型方法，可应用于自动化系统，实现高通量SNP检测。(C)2007 Elsevier B. V.保留所有权利。
Single-nucleotide polymorphisms (SNPs) are one-base variations in DNA sequence that can often be helpful when trying to find genes responsible for inherited diseases. In this paper, a microarray-based method for typing single nucleotide polymorphisms (SNPs) using solid-phase polymerase chain reaction (PCR) on magnetic nanoparticles (MNPs) was developed. One primer with biotin-label was captured by streptavidin coated magnetic nanoparticles (SA-MNPs), and PCR products were directly amplified on the surface of SA-MNPs in a 96-well plate. The samples were interrogated by hybridization with a pair of dual-color probes to determine SNP, and then genotype of each sample can be simultaneously identified by scanning the microarray printed with the denatured fluorescent probes. The C677T polymorphisms of methylenetetrahydrofolate reductase (MTHFR) gene from 126 samples were interrogated using this method. The results showed that three different genotypes were discriminated by three fluorescence patterns on the microarray. Without any purification and reduction procedure, and all reactions can be performed in the same vessel, this approach will be a simple and labor-saving method for SNP genotyping and can be applicable towards the automation system to achieve high-throughput SNP detection. (C) 2007 Elsevier B.V. All rights reserved.