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

该奖项是根据2009年《美国复苏和再投资法案》(Public Law 111-5)0846505SteinGenetic Information获得的，遗传信息是生命科学和医学快速发展的关键，但它仍然很难获得，因为它是由DNA分子上的大量微小碱基编码的。目前，只有细胞的生化机制可以很容易地区分这四个DNA碱基。这个职业项目寻求模仿生物学的精致敏感性，通过三种强大技术的组合从单个分子中提取遗传信息：首先，生物结合蛋白将沿着DNA分子连接到特定序列，并通过在那里创建物理凸起来标记其位置。其次，DNA分子将通过固态纳米孔--薄膜上的纳米级孔--电穿，该孔也将用作电子探测器。第三，DNA分子将使用光镊仪以可控的、稳定的速度引导通过纳米孔。当体积庞大的结合蛋白通过纳米孔时，它们将阻碍盐离子通过纳米孔的流动，并作为可测量的电流凹陷被检测到。这种识别DNA上特定位置的能力，再加上纳米级的位置控制和光镊子的皮牛顿力敏感性，将使对单分子的新的基础研究成为可能。课程将探讨纳米尺度的物理，包括热涨落的作用，以及DNA在纳米孔中的运动是连续的，还是以粘滑的方式进行。生物学问题，关于DNA-蛋白质相互作用的特异性和强度，也将被讨论。这个项目将帮助为潜在的变革性纳米孔技术奠定基础，能够高速从单个DNA分子中提取重要的遗传信息。这个综合职业项目的教育部分将向从小学到研究生院的所有级别的学生传达分子尺度上独特的生命物理。将特别关注普罗维登斯公立学校系统，该系统的重点是服务于在科学界人数很少的学生群体。将为小学科学计划和高中课外计划开发教育模块，高中生和科学教师都将通过暑期研究经验参与这一职业项目的技术方面。在大学一级，学生将继续受益于这个项目的基础设施，因为他们参与了研究，并从一门关于纳米尺度的生物和软凝聚物质物理的新课程中受益。这一跨学科研究和教育计划将为准备在高科技经济快速增长的领域就业的学生提供培训和丰富的机会。最后，该项目的技术目标可能会对生命科学产生广泛的影响，使研究人员能够更好地获得全基因组遗传信息和DNA-蛋白质相互作用，从而产生新的发现和生物医学应用。

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5)0846505SteinGenetic 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.