CAREER: Probing the Sequence and Dynamics of Single DNA Molecules Using Solid-State Nanopores, Optical Tweezers, and Binding Proteins

Brown University, Providence RI

Investigators

Abstract

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5) 0846505 Stein Genetic information holds a key to rapid progress in the life sciences and medicine, yet it remains difficult to obtain because it is encoded in a vast sequence of tiny bases along DNA molecules. At present, only the biochemical machinery of the cell can easily tell the four DNA bases apart. This CAREER project seeks to mimic biology's exquisite sensitivity by extracting genetic information from a single molecule through a combination of three powerful techniques: First, biological binding proteins will be attached to a specific sequence along DNA molecules, and mark its locations by creating a physical bulge there. Second, the DNA molecule will be electrically threaded through a solid-state nanopore - a nanometer-scale hole in a thin membrane - that will also serve as an electrical detector. Third, the DNA molecule will be guided through the nanopore at a controlled, steady speed using an optical tweezers instrument. As the bulky binding proteins pass through the nanopore, they will impede the flow of salt ions through the nanopore, and be detected as a measureable dip in electrical current. This ability to identify specific locations along DNA, combined with the nanometer-scale position control and pico-Newton force sensitivity of optical tweezers, will enable new fundamental studies on single molecules. Nanoscale physics will be probed, including the role of thermal fluctuations, and whether the motion of DNA through a nanopore is continuous, or proceeds in a stick-slip manner. Biological questions, concerning the specificity and strength of DNA-protein interactions, will also be addressed. This project will help lay the groundwork for a potentially transformative nanopore technology, capable of extracting important genetic information from single DNA molecules at high speed. The educational component of this integrated CAREER project will convey the unique physics of life at the molecular scale to students at all levels from elementary school through graduate school. Particular attention will be focused on the Providence Public School system, which is stressed, and which serves a community of students that are largely underrepresented in the sciences. Education modules will be developed for both elementary school science programs and high school after-school programs, and both high school students and science teachers will participate in the technical side of this CAREER project through summer research experiences. At the university level, students will continue to benefit from the infrastructure of this project as they participate in research, and from a new course on the physics of biological and soft condensed matter at the nanoscale. This interdisciplinary research and education program will provide training and enrichment opportunities for students preparing for careers in a rapidly growing area of the high-technology economy. Finally, the technological objectives of this project may have a broad impact on the life sciences by giving researchers improved access to genome-wide genetic information and DNA-protein interactions, from which new discoveries and biomedical applications can derive.