Modeling Macromolecular Transport for Sequencing Technologies

描述(由申请人提供)：迫切需要开发革命性的技术，以快速和经济地对大DNA分子进行测序，这导致了许多实验策略。其中最主要的是基于纳米孔的电泳实验。在这些实验中，单分子DNA在外加电场下通过蛋白质通道和固态纳米孔时的移位被监测。虽然这些实验的结果非常有希望达到1000美元的基因组目标，但仍有许多谜题，这些纳米系统的物理需要从基本的科学角度来理解。这项拟议的研究涉及对纳米孔环境中DNA在电动力和流体动力影响下的行为的基本理解。我们将调查几个关键系统组件背后的挑战，目标是将哺乳动物大小的基因组测序成本降低到1000美元。主要的挑战涉及在纳米孔处捕获目标分子的可预测性、有效地穿入孔中以及减缓分子通过孔的转移。我们将结合统计力学理论、计算机模拟和耦合非线性方程的数值计算来处理DNA移位现象中的聚合物统计和动力学、静电学和流体动力学。虽然拟议的研究一般与所有基于纳米孔的实验有关，但将具体取决于：(A)杂交在通过a-溶血素、MSPA和固态孔转位中的作用，(B)通过通道的酶调节DNA转位，以及(C)控制DNA在蛋白质通道和固态纳米孔中的捕获率和转位成功率。 与公共卫生相关：可获得低成本的DNA测序技术对于确定疾病来源和维护公共健康至关重要。这项拟议的研究解决了基因组测序技术开发中几个关键系统组件的挑战，每个哺乳动物大小的基因组的成本为1000美元。

DESCRIPTION (provided by applicant): The urgent need to develop revolutionary technologies, for sequencing large DNA molecules quickly and economically, has led to many experimental strategies. Chief among these are the nanopore-based electrophoretic experiments. In these experiments, translocation of single molecules of DNA is monitored as they pass through protein channels and solid-state nanopores under an external electric field. While the results from such experiments are extremely promising towards reaching $1000 genome target, there are many puzzles and the physics of these nanoscopic systems needs to be understood from a fundamental scientific point of view. The proposed research deals with a fundamental understanding of the behavior of DNA in nanopore environments under the influence of electrical and hydrodynamic forces. We will investigate the challenges underlying several key system components in the goal of reducing the cost of sequencing mammalian-sized genomes to $1000. The major challenges deal with the predictability of capture of the target molecule at the nanopore, efficient threading into the pore, and slowing down the translocating molecule through the pore. We will use a combination of statistical mechanics theory, computer simulations, and numerical computation of coupled nonlinear equations to address polymer statistics and dynamics, electrostatics, and hydrodynamics in the phenomena of DNA translocation. The proposed research, while being generally relevant to all nanopore-based experiments, will be hinged specifically on: (a) role of hybridization in translocation through a-hemolysin, MspA, and solid-state pores, (b) enzyme-modulated DNA translocation through channels, and (c) control of capture rate and successful translocation rate of DNA in protein channels and solid-state nanopores. PUBLIC HEALTH RELEVANCE: Availability of low-cost technologies for DNA sequencing is vital in identifying the origins of diseases and maintenance of public health. The proposed research addresses the challenges in several key system components in the development of genome sequencing technologies at the cost of $1000 per a mammalian-sized genome.