Clustered Regularly Interspaced Short Palindromic Repeats/Cas9-Mediated Lateral Flow Nucleic Acid Assay
Clustered Regularly Interspaced Short Palindromic Repeats/Cas9-Mediated Lateral Flow Nucleic Acid Assay
复制标题
成簇规则间隔短回文重复序列/Cas9介导的侧流核酸测定
DOI：
10.1021/acsnano.0c00022
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发表时间：
2020-02-01
期刊：
影响因子：
17.1
通讯作者：
Zhou, Xiaoming
中科院分区：
文献类型：
--
作者：
Wang, Xusheng;Xiong, Erhu;Zhou, Xiaoming
横向流动分析是分析免疫应答最方便的分析技术之一，但其在精确遗传分析中的适用性受到假阳性信号和繁琐低效的杂交步骤的限制。在这里，我们将CRISPR（聚集规律间隔短回文重复序列）/Cas系统引入到侧流分析中，称为CRISPR/ cas9介导的侧流核酸测定（CASLFA），以解决这些问题。在本研究中，CASLFA被用于鉴定单核细胞增生李斯特菌、转基因生物（GMOs）和非洲猪瘟病毒（ASFV），在1小时内的检测下限为数百个基因组样本，具有高特异性。我们进一步评估了CASLFA在非实验室环境中的性能，并成功地从110份疑似猪血清样本中确认了27份ASFV感染样本，与实时荧光定量PCR （RT-PCR）相比，准确度为100%。由于其快速和准确，CASLFA满足了下一代分子诊断工具的一些特征，允许在不需要技术专业知识和复杂辅助设备的情况下进行即时使用。这种方法在资源贫乏或非实验室环境中具有很大的基因分析潜力。
The lateral flow assay is one of the most convenient analytical techniques for analyzing the immune response, but its applicability to precise genetic analyses is limited by the false-positive signal and tedious and inefficient hybridization steps. Here, we introduce the CRISPR (clustered regularly interspaced short palindromic repeats) /Cas system into the lateral flow assay, termed CRISPR/Cas9-mediated lateral flow nucleic acid assay (CASLFA), to address such issues. In this study, CASLFA is utilized to identify Listeria monocytogenes, genetically modified organisms (GMOs), and African swine fever virus (ASFV) at a detection limit of hundreds of copies of genome samples with high specificity within 1 h. We further evaluated the performance of CASLFA in a nonlaboratory environment and successfully confirmed 27 ASFV-infected samples from 110 suspected swine serum samples, with an accuracy of 100% when compared to real-time PCR (RT-PCR) assay. CASLFA satisfies some of the characteristics of a next-generation molecular diagnostics tool due to its rapidity and accuracy, allowing for point-of-care use without the need for technical expertise and complex ancillary equipment. This method has great potential for gene analysis in resource-poor or nonlaboratory environments.