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，当DNA的单分子在外部电场下通过蛋白质通道和固态纳米孔时，监测它们的易位。 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，但仍存在许多难题，需要从基础科学的角度来理解这些纳米系统的物理学。 拟议的研究涉及对DNA在纳米孔环境中受电力和水动力影响的行为的基本理解。 我们将研究几个关键系统组件的挑战，以降低测序的成本为目标，以1000美元为单位的基因组。 主要的挑战涉及在纳米孔处捕获靶分子的可预测性、有效地穿入孔中以及减缓易位分子通过孔。 我们将使用统计力学理论，计算机模拟和耦合非线性方程的数值计算相结合，以解决DNA易位现象中的聚合物统计和动力学，静电学和流体力学。 所提出的研究虽然通常与所有基于纳米孔的实验相关，但将具体取决于：（a）杂交在通过α-溶血素、MspA和固态孔的易位中的作用，（B）通过通道的酶调节的DNA易位，以及（c）在蛋白质通道和固态纳米孔中DNA的捕获率和成功易位率的控制。 公共卫生相关性：提供低成本的DNA测序技术对于确定疾病的起源和维护公共卫生至关重要。 拟议的研究解决了在基因组测序技术的发展中的几个关键系统组件的挑战，每1000美元的成本为一个大的基因组。

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.