Single-Molecule Imaging of Biorecognition Kinetics

University Of Utah, Salt Lake City UT

Investigators

Abstract

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.