Wang Lin, Liu Zhanmin, Xia Xueying, Yang Cuiyun, Huang Junyi, Wan Sibao
School of Life Sciences, Shanghai University, Shanghai, 200444, China.
School of Life Sciences, Shanghai University, Shanghai, 200444, China.
J Virol Methods. 2017 May;243:113-119. doi: 10.1016/j.jviromet.2017.01.010. Epub 2017 Jan 18.
Cucumber green mottle mosaic virus (CGMMV)causes a severe mosaic symptom of watermelon and cucumber, and can be transmitted via infected cucumber seeds, leaves and soil. It remains a challenge to detect this virus to prevent its introduction and infection and spread in fields. For this purpose, a simple and sensitive label-free colorimetric detection method for CGMMV has been developed with unmodified gold nanoparticles (AuNPs) as colorimetric probes. The method is based on the finding that the presence of RT-PCR target products of CGMMV and species-specific probes results in color change of AuNPs from red to blue after NaCl induction. Normally, species-specific probes attach to the surface of AuNPs and thereby increasing their resistance to NaCl-induced aggregation. The concentration of sodium, probes in the reaction system and evaluation of specificity and sensitivity of a novel assay, visual detection of Cucumber green mottle mosaic virus using unmodified AuNPs has been carried out with simple preparation of samples in our study. Through this assay, as low as 30pg/μL of CGMMV RNA was thus detected visually, by the naked eye, without the need for any sophisticated, expensive instrumentation and biochemical reagents. The specificity was 100% and exhibited good reproducibility in our assays. The results note that this assay is highly species-specific, simple, low-cost, and visual for easy detection of CGMMV in plant tissues. Therefore, visual assay is a potentially useful tool for middle or small-scales corporations and entry-exit inspection and quarantine bureau to detect CGMMV in cucumber seeds or plant tissues.
黄瓜绿斑驳花叶病毒（CGMMV）会导致西瓜和黄瓜出现严重的花叶症状，并且可通过受感染的黄瓜种子、叶片和土壤传播。检测这种病毒以防止其传入、感染并在田间传播仍然是一项挑战。为此，已开发出一种简单且灵敏的无标记比色检测方法来检测CGMMV，该方法使用未修饰的金纳米颗粒（AuNPs）作为比色探针。该方法基于以下发现：CGMMV的逆转录聚合酶链反应（RT-PCR）靶标产物和物种特异性探针的存在会导致在氯化钠诱导后AuNPs的颜色从红色变为蓝色。通常，物种特异性探针附着在AuNPs表面，从而增加其对氯化钠诱导聚集的抗性。在我们的研究中，通过简单的样品制备，对反应体系中的钠浓度、探针以及一种新型检测方法的特异性和灵敏度进行了评估，即使用未修饰的AuNPs对黄瓜绿斑驳花叶病毒进行可视化检测。通过该检测方法，肉眼即可直观检测到低至30pg/μL的CGMMV RNA，无需任何复杂、昂贵的仪器和生化试剂。在我们的检测中，特异性为100%，且具有良好的重现性。结果表明，该检测方法具有高度的物种特异性、简单、低成本且可视化，便于在植物组织中检测CGMMV。因此，可视化检测方法对于中、小型企业以及出入境检验检疫局检测黄瓜种子或植物组织中的CGMMV而言是一种潜在的有用工具。
