Gold nanoparticle-mediated nucleic acid isothermal amplification with enhanced specificity
Gold nanoparticle-mediated nucleic acid isothermal amplification with enhanced specificity
复制标题
金纳米颗粒介导的核酸等温扩增具有增强的特异性
DOI：
10.1016/j.aca.2018.09.016
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发表时间：
2018
影响因子：
6.2
通讯作者：
Jilie Kong
中科院分区：
文献类型：
--
作者：
Xin Ye;Xueen Fang;Xinxin Li;Jilie Kong
环介导等温扩增技术是一种很有前途的核酸检测方法。然而，它具有高的假阳性扩增率，这在很大程度上限制了它的应用。在本研究中，我们观察到金纳米粒子（AuNP）吸附单链DNA引物，并通过静电吸附与BstDNA聚合酶相互作用。作为这些相互作用的结果，金纳米颗粒的存在对环介导的等温扩增系统产生热启动效应。基于这些结果，我们开发了一种新的金纳米粒子介导的核酸等温扩增检测。该方法的特异性显著提高，在48 °C的热启动温度下，轮状病毒和β-actin基因的假阳性率分别从76%和100%降至0%。与传统方法相比，该方法具有较高的灵敏度和较好的检测限（1 × 103 copies/μL）。这种方法有可能解决环介导等温扩增的非特异性问题，从而促进其在现实世界中的应用，特别是在临床环境中。
Loop-mediated isothermal amplification is a promising method in the area of nucleic acid detection. However, it suffers from a high rate of false-positive amplifications that largely restrict its application. In this study, we observed gold nanoparticles (AuNP) absorbing single-stranded DNA primers and interacting withBstDNA polymerase via electrostatic adsorption. As a result of these interactions, the presence of the gold nanoparticles exerted a hot-start effect on the loop-mediated isothermal amplification system. Based on these results, we developed a novel AuNP-mediated nucleic acid isothermal amplification assay. This assay displays significantly enhanced specificity—the proportion of false positive decreased from 76% to 0% and from 100% to 0% for the detection of rotavirus and the β-actin gene, respectively, with the hot-start temperature of 48 °C. Moreover, our AuNP-based assay maintained good sensitivity and a satisfactory detection limit (1 × 103copies/μL) compared with the conventional assay. This approach has the potential to solve the nonspecificity problem of loop-mediated isothermal amplification, thereby promoting its real-world application, particularly, in clinical settings.