使用双纳米孔装置区分dsDNA构建体上的蛋白质标签
Discriminating protein tags on dsDNA constructs using a dual Nanopore device
摘要：
我们报告了一种新的模拟策略，使我们能够确定关键参数，控制实验测量的特性的结构蛋白质标签上的dsDNA构建易位通过双纳米孔设置。首先，我们通过复制和解释链霉亲和素标记物在48 kbp长的dsDNA上的实验停留时间分布的物理起源来验证该方案。这些研究揭示了与dsDNA片段的动力学相比，蛋白质标签特征的重要差异，立即提供了如何改进测量方案以准确破译未知基因组长度的线索。特别重要的是，在硅片上的电场的影响，内部和外部的孔，我们发现是关键的区别蛋白质标签的基础上，通过一个通用的幂律依赖性的平均停留时间在每个孔揭示了它们的有效电荷和质量。模拟协议能够监测分段动态的单个单体在亚纳米长度尺度，并提供了不同的速度变化的解释从一个标签到其他使用非平衡张力传播理论，一个关键因素，以准确地破译基因组长度。通过计算与实验结果吻合较好的Peclet数，进一步验证了模型和模拟参数的合理性。从CG模型对我们的模拟结果的分析具有改进实验获得的基因组长度的准确性的能力，并且仔细选择的模拟策略可以作为根据其物理特征来区分dsDNA构建体上不同来源的不同类型的中性和带电标签的有力工具，并且可以提供见解以提高实验双重标记的效率和准确性。纳米孔设置。
英文摘要：
We report a novel simulation strategy that enables us to identify key parameters controlling the experimentally measurable characteristics of structural protein tags on dsDNA construct translocating through a double nanopore setup. First, we validate the scheme in silico by reproducing and explaining the physical origin of the experimental dwell time distributions of the Streptavidin markers on a 48 kbp long dsDNA. These studies reveal the important differences in the characteristics of the protein tags compared to the dynamics of dsDNA segments, immediately providing clues on how to improve the measurement protocols to decipher the unknown genomic lengths accurately. Of particular importance is the in silico studies on the effect of electric field inside and beyond the pores which we find is critical to discriminate protein tags based on their effective charges and masses revealed through a generic power-law dependence of the average dwell time at each pore. The simulation protocols enable to monitor piecewise dynamics of the individual monomers at a sub-nanometer length scale and provide an explanation of the disparate velocity variation from one tag to the other using the nonequilibrium tension propagation theory, - a key element to decipher genomic lengths accurately. We further justify the model and the chosen simulation parameters by calculating the Peclet number which is in close agreement with the experiment. Analysis of our simulation results from the CG model has the capability to refine the accuracy of the experimentally obtained genomic lengths and carefully chosen simulation strategies can serve as a powerful tool to discriminate different types of neutral and charged tags of different origins on a dsDNA construct in terms of their physical characteristics and can provide insights to increase both the efficiency and accuracy of an experimental dual-nanopore setup.
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