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。这些不同的和重要的功能促使传感器的发展，允许灵敏的检测和识别的RNA分子。该项目将为两种特定类别的RNA分子开发RNA传感器：i.微RNA（miRNA），具有作为疾病如癌症的生物标志物的潜力的调节类RNA，和ii.）病毒RNA，以便能够快速检测RNA病毒，如冠状病毒（Covid-19）。所提出的传感器平台包含一个DNA链，其末端连接到两个金属纳米颗粒，其光学信号取决于它们的分离。DNA传感器链被设计成当它与特定的RNA靶结合时改变其结构。这种结构修饰改变了纳米颗粒探针之间的间隔，并改变了它们的信号，即使对于小的纳米颗粒，也可以通过散射信号与参考光束的干涉灵敏地检测到它们。该传感器将被设计为直接检测miRNA，但对于病毒RNA，耦合的纳米颗粒传感器将与酶信号放大步骤相结合，以获得灵敏且快速可重构的检测平台。这项研究将被纳入教育和推广活动，包括课程工作的发展，以支持研究，组织一个夏季研讨会的基本原理等离子体和纳米科学的内城高中学生在波士顿地区和他们的老师，旨在激发他们对科学和技术的职业生涯，以及开发一个Instagram频道，为教育非专业观众了解等离子体纳米传感器提供一个很好的平台。该项目将结合联合收割机分子信标，如DNA结构，其末端用贵金属纳米颗粒（NP）功能化。探针与双色干涉散射显微镜（iSCAT）检测，以产生用于两种重要类别的RNA的基于等离子体耦合的传感器：i.）调节性miRNA和ii.）严重急性呼吸综合征冠状病毒2（SARS-CoV-2）RNA。靶RNA与传感器的结合诱导了从“闭合”构象到“开放”构象的构象变化，并改变了与DNA链结合的NP标记之间的等离子体偶联。双色iSCAT显微镜将被应用于提高检测与传感器结合到其目标时的开口相关的干涉散射信号变化的灵敏度。传感器的DNA识别元件将被设计为直接与miRNA结合，而对于病毒RNA，等离子体换能器将与CRISPR-CAS 13检测和随后的样品扩增相结合，以获得灵敏且可重构的病毒RNA检测平台。该分子信标不直接与病毒RNA结合，而是识别通过CAS 13反式切割底物产生的RNA片段。该奖项反映了NSF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.