Single-Molecule Imaging of Biorecognition Kinetics

生物识别动力学的单分子成像

• 批准号: 1608949
• 负责人: Joel Harris
• 金额: $ 52.82万
• 依托单位: University of Utah
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2020-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1608949&HistoricalAwards=false
• 关键词: Single; Molecule; Imaging; Biorecognition; Kinetics

The Chemical Measurement and Imaging (CMI) program of the Division of Chemistry at the National Science Foundation supports this project. Professor Joel M. Harris and his research group at the University of Utah are developing microscopy experiments to investigate reactions of biological molecules, especially DNA. The chemical reaction between complementary strands of DNA is the fundamental step in duplication and transcription of genetic information in living organisms. The goal of this project is to develop microscopy methods that allow the reactions between DNA strands to be observed in "movies" of individual molecules as they react at a glass surface. Understanding these reactions is critical to the development of DNA-based genetic testing used in disease diagnosis. Single-molecule imaging also serves as a sensitive detection method, where counting individual molecules enables the analysis of extremely low concentrations of target DNA in biological specimens. The broader impacts are demonstrated through the training of graduate students, undergraduates, and postdoctoral associates in microscopy and bioanalytical chemistry. These topics are relevant to advancing U.S. science and technology. The project also supports K-12 science education and outreach through development of Chemistry-under-the-Microscope exercises for middle- and high-school students.The unique approach taken in this project is to immobilize unlabeled DNA on a capture surface that is engineered to have extremely low non-specific interactions with fluorescently-labeled target DNA in solution. The result is a high sensitivity measurement platform, where capture surfaces can be used for long periods of time, and even reused in multiple experiments. With super-resolution imaging, individual immobilized DNA probe strands on the surface can be identified, thereby creating a microarray of single-molecule capture sites. Impacts of the research include generating understanding of structural factors that influence the performance of DNA-based biosensors. The development of microscopy instrumentation for single-molecule imaging supports several collaborations including DNA aptamer discovery, measuring membrane affinity of signaling proteins, and assessing of new drug candidates.

美国国家科学基金会化学部的化学测量和成像（CMI）项目支持该项目。犹他大学的 Joel M. Harris 教授和他的研究小组正在开发显微镜实验来研究生物分子（尤其是 DNA）的反应。 DNA 互补链之间的化学反应是生物体中遗传信息复制和转录的基本步骤。该项目的目标是开发显微镜方法，允许在玻璃表面反应时在单个分子的“电影”中观察 DNA 链之间的反应。了解这些反应对于疾病诊断中基于 DNA 的基因检测的发展至关重要。单分子成像还可以作为一种灵敏的检测方法，通过对单个分子进行计数，可以分析生物样本中极低浓度的目标 DNA。通过对研究生、本科生和博士后进行显微镜和生物分析化学方面的培训，证明了更广泛的影响。这些主题与推进美国科学技术相关。该项目还通过为中学生和高中生开发显微镜下化学练习来支持 K-12 科学教育和推广。该项目采用的独特方法是将未标记的 DNA 固定在捕获表面上，该表面经过设计，与溶液中荧光标记的目标 DNA 具有极低的非特异性相互作用。结果是一个高灵敏度的测量平台，捕获表面可以长期使用，甚至可以在多个实验中重复使用。通过超分辨率成像，可以识别表面上单个固定的 DNA 探针链，从而创建单分子捕获位点的微阵列。 该研究的影响包括加深对影响 DNA 生物传感器性能的结构因素的理解。用于单分子成像的显微镜仪器的开发支持了多项合作，包括 DNA 适体发现、测量信号蛋白的膜亲和力以及评估新候选药物。