Detection of single-nucleotide polymorphisms with novel leaky surface acoustic wave biosensors, DNA ligation and enzymatic signal amplification
Detection of single-nucleotide polymorphisms with novel leaky surface acoustic wave biosensors, DNA ligation and enzymatic signal amplification
复制标题
利用新型渗漏表面声波生物传感器、DNA 连接和酶信号放大检测单核苷酸多态性
DOI：
10.1016/j.bios.2011.12.036
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发表时间：
2012-03-15
影响因子：
12.6
通讯作者：
Chen, Ming
中科院分区：
文献类型：
--
作者：
Xu, Qinghua;Chang, Kai;Chen, Ming
This manuscript describes a new technique for detecting single-nucleotide polymorphisms (SNPs) by integrating a leaky surface acoustic wave (LSAW) biosensor, enzymatic DNA ligation and enzymatic signal amplification. In this technique, the DNA target is hybridized with a capture probe immobilized on the surface of a LSAW biosensor. Then, the hybridized sequence is ligated to biotinylated allele-specific detection probe using Taq DNA ligase. The ligation does not take place if there is a single-nucleotide mismatch between the target and the capture probe. The ligated detection probe is transformed into a streptavidin-horseradish peroxidase (SA-HRP) terminal group via a biotin-streptavidin complex. Then, the SA-HRP group catalyzes the polymerization of 3,3-diaminobenzidine (DAB) to form a surface precipitate, thus effectively increasing the sensitivity of detecting surface mass changes and allowing detection of SNPs. Optimal detection conditions were found to be: 0.3 mol/L sodium ion concentration in PBS, pH 7.6, capture probe concentration 0.5 mu mol/L and target sequence concentration 1.0 mu mol/L. The detection limit was found to be 1 x 10(-12) mol/L. Using this technique, we were able to detect a single-point mutation at nucleotide A2293G in Japanese encephalitis virus. (C) 2011 Elsevier B.V. All rights reserved.
