RAPID: Enzyme-free detection of coronavirus RNA using reconfigurable DNA nanoswitches

RAPID：使用可重构 DNA 纳米开关无酶检测冠状病毒 RNA

• 批准号: 2030279
• 负责人: Ken Halvorsen
• 金额: $ 17.78万
• 依托单位: SUNY at Albany
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2021-04-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2030279&HistoricalAwards=false
• 关键词: RAPID; Enzyme; free; detection; coronavirus

A critical aspect of controlling the current COVID-19 outbreaks is early and fast detection of the infected population. Current methods for detecting the novel coronavirus that causes COVID-19 typically require a laboratory and use expensive equipment and reagents. These requirements limit the number of tests that can be performed nationwide. In this project, the research team aims to develop an alternative diagnostic approach that directly detects the viral RNA without any amplification, labeling, or the use of enzymes. The assay requires minimal steps and uses a compact, bufferless gel cartridge system that can be deployed in a clinical setting. Beyond the current COVID-19 outbreak, this sensing platform can also apply to other existing or emerging viruses.This work aims to develop a nanoswitch assay sensitive enough to detect clinical levels of SARS-CoV-2 viral RNA within 1 hour. Once the viral RNA is extracted from a sample, the RNA is fragmented and mixed with the DNA-based nanoswitches. Hybridization of a viral RNA fragment of a specific sequence with a nanoswitch induces a conformational change of the engineered DNA strand from its linear "off" state to a looped "on" state. After separation of the target-bound looped form from the linear nanoswitches by electrophoresis in a gel cartridge system, the viral load is quantified from the intensity of the looped (on) nanoswitches in a gel image.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

控制当前COVID-19疫情的一个关键方面是早期和快速发现受感染人群。目前检测导致COVID-19的新型冠状病毒的方法通常需要实验室，并使用昂贵的设备和试剂。这些要求限制了可以在全国范围内进行的测试的数量。在这个项目中，研究小组的目标是开发一种替代的诊断方法，直接检测病毒RNA，而不需要任何扩增、标记或使用酶。该分析需要最少的步骤，并使用一个紧凑的，无缓冲凝胶盒系统，可以部署在临床设置。除了当前的COVID-19疫情，该传感平台还可以应用于其他现有或新出现的病毒。这项工作旨在开发一种足够灵敏的纳米开关试验，以在1小时内检测出SARS-CoV-2病毒RNA的临床水平。一旦从样本中提取出病毒RNA， RNA就会被分割并与基于dna的纳米开关混合。将特定序列的病毒RNA片段与纳米开关杂交，诱导工程DNA链的构象变化，从其线性“关闭”状态变为环状“打开”状态。在凝胶盒系统中通过电泳将目标结合的环状形式从线性纳米开关中分离出来后，通过凝胶图像中环状（打开）纳米开关的强度来定量病毒载量。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。