Next Generation Plasmon Coupling Nanosensors

核糖核酸（RNA）分子在控制基因表达和控制细胞行为方面具有多种功能。RNA也被一些病毒用来储存遗传信息。这些多样而重要的功能促使传感器的发展，允许一个敏感的检测和识别RNA分子。该项目将为两种特定类型的RNA分子开发RNA传感器：1)microRNA (miRNA)，这是一种具有作为癌症等疾病生物标志物潜力的调控类RNA； 2)病毒RNA，能够快速检测RNA病毒，如冠状病毒（Covid-19）。所提出的传感器平台包含一条DNA链，其两端连接到两个金属纳米颗粒，其光学信号依赖于它们的分离。DNA传感器链被设计成在与特定RNA目标结合时改变其结构。这种结构的改变改变了纳米颗粒探针之间的分离，改变了它们的信号，即使是小的纳米颗粒，也可以通过参考光束干扰散射信号来敏感地检测到。该传感器将被设计用于直接检测miRNA，但对于病毒RNA，耦合纳米颗粒传感器将与酶信号扩增步骤相结合，以获得灵敏且快速可重构的检测平台。研究将整合到教育和推广活动中，包括开发课程作业以支持研究，为波士顿地区内城高中学生和他们的老师组织一个关于等离子体和纳米科学基础的暑期研讨会，旨在激发他们对科学和技术事业的兴趣。并开发一个Instagram频道，为外行提供一个很好的平台，让他们了解等离子纳米传感器。该项目将结合分子信标式的DNA结构，其末端被贵金属纳米颗粒（NP）探针功能化，并结合双色干涉散射显微镜（iSCAT）检测，为两类重要的RNA生成基于等离子体耦合的传感器：i.)调性miRNA和ii.)严重急性呼吸综合征冠状病毒2 (SARS-CoV-2) RNA。与传感器结合的靶RNA诱导构象从“封闭”到“开放”的变化，并改变与DNA链结合的NP标签之间的等离子体耦合。将应用双色iSCAT显微镜来提高检测与传感器与目标结合时打开相关的干涉散射信号变化的灵敏度。传感器的DNA识别元件将被设计成直接与miRNA结合，而对于病毒RNA，等离子体换能器将与CRISPR-CAS13检测和随后的样品扩增相结合，以获得一个敏感且可重构的病毒RNA检测平台。分子信标不直接与病毒RNA结合，而是识别CAS13反式切割底物产生的RNA片段。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。

Ribonucleic acid (RNA) molecules have diverse functions in controlling gene expression and controlling cellular behavior. RNA is also used by some viruses to store genetic information. These diverse and important functions motivate the development of sensors that allow for a sensitive detection and identification of RNA molecules. This project will develop RNA sensors for two specific classes of RNA molecules: i.) microRNA (miRNA), a regulatory class of RNA that has potential as biomarkers for diseases such as cancer, and ii.) viral RNA to enable a rapid detection of RNA viruses, such as the corona virus (Covid-19). The proposed sensor platform contains a DNA strand connected at its ends to two metal nanoparticles whose optical signal depend on their separation. The DNA sensor strand is designed to change its structure when it binds to a specific RNA target. This structural modification changes the separation between the nanoparticle probes and alters their signal, which is sensitively detected even for small nanoparticles via interference of the scattered signal with a reference light beam. The sensor will be designed to detect miRNA directly, but for viral RNA the coupled nanoparticle sensor will be combined with an enzymatic signal amplification step to obtain a sensitive and rapidly reconfigurable detection platform. The research will be integrated into education and outreach activities that include development of course work to support the research, organization of a summer workshop on the fundamentals of plasmonics and nanoscience for inner city high school students in the Boston area and their teachers that is designed to excite them about a career in science and technology, and development of an Instagram channel that will provide an excellent platform for educating a lay audience about plasmonic nanosensors.The project will combine molecular beacon like DNA structures whose ends are functionalized with noble metal nanoparticle (NP) probes with two-color interferometric scattering microscopy (iSCAT) detection to generate a plasmon coupling based sensor for two important classes of RNA: i.) regulatory miRNA and ii.) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA. Target RNA binding to the sensor induces a conformational change from a “closed” to an “open” conformation and alters the plasmon coupling between the NP labels bound to the DNA strand. Two-color iSCAT microscopy will be applied to enhance the sensitivity for detecting changes in the interferometric scattering signal associated with the opening of the sensor upon binding to its target. The DNA recognition element of the sensor will be designed to bind to miRNA directly, while for the viral RNA the plasmonic transducer will be combined with CRISPR-CAS13 detection and subsequent sample amplification to obtain a sensitive and reconfigurable viral RNA detection platform. Instead of directly binding to viral RNA, the molecular beacon will recognize RNA fragments generated through CAS13 trans-cleavage of a substrate.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.