Mathematical Modeling of a Self-Assembled Probe for Non Intercalator Type Real-Time Monitoring of PCR

用于非嵌入剂型 PCR 实时监测的自组装探针的数学建模

• 批准号: 9813297
• 负责人: Fereshteh Emami
• 金额: $ 34.66万
• 依托单位: SOUTHEASTERN LOUISIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9813297
• 关键词: Acids; Address; Algorithms; Attention; Behavior; Binding; Biochemistry; Biological; Biosensing Techniques; Buffers; Catechols; Chemicals; Clinical Research; Complex; Computer Simulation; Consumption; DNA; DNA Probes; DNA Sequence; DNA amplification; Data; Detection; Diagnostics Research; Diphosphates; Drug Screening; Dyes; Equilibrium; Experimental Designs; Fluorescence; GC Rich Sequence; Goals; Growth; Imagery; Intercalating Agents; Ionic Strengths; Knowledge; Laws; Measurement; Medical; Methods; Modeling; Molecular; Monitor; Organism; Outcome; Pathway interactions; Pharmacologic Substance; Polymerase Chain Reaction; Process; Reaction; Research; Resolution; Ribonucleotides; Salts; Scheme; Science; Screening procedure; Signal Transduction; Structure; System; Technology; Temperature; Testing; Thermodynamics; Time; Vertebral column; aptamer; aqueous; base; chemical reaction; computational basis; innovation; inorganic phosphate; insight; mathematical algorithm; mathematical model; melting; molecular diagnostics; molecular modeling; novel; polymerization; real time monitoring; self assembly; sensor; simulation; time use; tool; tripolyphosphate

PROJECT SUMMARY/ABSTRACT Real-time PCR has become a crucial tool in many fields of molecular diagnostics, and many methods have been developed to monitor PCR as the reaction proceeds. The objective of this application is to examine sensing molecules such as the assembles of ZnII–DPA-attached phenylboronic acid (1.Zn) and catechol-type dyes like alizarin red S (ARS) for non-intercalator type real-time monitoring of PCR using mathematical modeling and experimental observation. The central hypothesis, which was formulated based on preliminary data, is that the 1.Zn–ARS sensor is selective towards pyrophosphate (PPi), the byproduct of DNA amplification, over the other phosphates. Different analytical evidence displayed existence multiple intertwined equilibria for the self- assembled 1.Zn-dyes sensors, yet the molecular mechanisms that underlie the phosphate sensing processes are not well understood. The long-term goal is to understand the molecular details of such very complicated self- assembly combinations that are not only limited to these examples through mathematical modeling, and to broaden this knowledge by developing novel biosensing strategies. The rationale for the proposed research is to identify unknown influences on ribonucleotide detection processes, to advance our understanding of self- assembled supramolecular host-guest thermodynamics, and to generate testable hypotheses for aptamer and drug screening procedures. To address these goals, we aim to develop a mathematical algorithm that can model the entire twenty-seven molecular intertwined interactions and reactions among the supramolecular probe, 1.Zn- ARS, and biological phosphates. We will determine the thermodynamic parameters of the interactions and reactions using the developed mathematical model, experimental design, and data fitting of the potentiometric measurements. We will then introduce optimum experimental conditions for the best real-time oligophosphate monitoring based on mathematical modeling and computer simulations. In addition, as a proof of principle, we will validate the simulated results with the experimental real-time PCR observations. This study is innovative because a sophisticated mathematical model and computer simulations will be used to investigate the reaction pathways for the formation of different guest-host bindings of 1.Zn-ARS-PPi. The computer simulations will enable the visualization of the effects of different values of binding constants, temperatures, ionic strengths, concentrations, starting pHs, etc. for any possible experimental conditions and provide appropriate directions for real-time biomolecular sensing. The proposed project is significant because, by combining the power of supramolecular self-assemblies with mathematical modeling, a better understanding of 1.Zn-ARS-PPi molecular interactions within their complex chemical networks will be achieved that can be generalized to many other complex chemical processes in pharmaceutical and biochemistry sciences to help detect and monitor analytes of medical importance.

项目概要/摘要 实时荧光定量PCR已成为分子诊断许多领域的重要工具，许多方法已被广泛应用。 开发用于监测反应过程中的 PCR。该应用的目的是检查传感 分子，例如 ZnII-DPA 连接的苯基硼酸 (1.Zn) 和儿茶酚型染料的组装体 茜素红 S (ARS)，用于使用数学模型和非嵌入剂型实时监测 PCR 实验观察。根据初步数据提出的中心假设是 1.Zn-ARS传感器对DNA扩增副产物焦磷酸盐（PPi）具有选择性，优于其他传感器 磷酸盐。不同的分析证据表明，自我平衡存在多重交织的平衡。 组装1.锌染料传感器，但磷酸盐传感过程的分子机制 没有被很好地理解。长期目标是了解这种非常复杂的自体分子的细节 通过数学建模的装配组合不仅限于这些示例，而且还可以 通过开发新颖的生物传感策略来拓宽这一知识。拟议研究的理由是 识别对核糖核苷酸检测过程的未知影响，增进我们对自我的理解 组装超分子主客体热力学，并生成适体和可测试的假设 药物筛选程序。为了实现这些目标，我们的目标是开发一种可以建模的数学算法 超分子探针1.Zn-之间全部27个分子相互交织的相互作用和反应 ARS 和生物磷酸盐。我们将确定相互作用的热力学参数和 使用开发的数学模型、实验设计和电位计数据拟合进行反应 测量。然后我们将介绍最佳实时低聚磷酸盐的最佳实验条件 基于数学建模和计算机模拟的监测。此外，作为原理证明，我们 将通过实验实时 PCR 观察来验证模拟结果。这项研究具有创新性 因为将使用复杂的数学模型和计算机模拟来研究反应 1.Zn-ARS-PPi 不同客体-主体结合形成的途径。计算机模拟将 能够可视化结合常数、温度、离子强度的不同值的影响， 浓度、起始 pH 值等任何可能的实验条件，并提供适当的指导 实时生物分子传感。拟议的项目意义重大，因为通过结合各方的力量 1.Zn-ARS-PPi分子超分子自组装与数学建模，更好的理解 他们复杂的化学网络内的相互作用将被实现，并且可以推广到许多其他 制药和生物化学科学中的复杂化学过程，有助于检测和监测分析物 具有医学重要性。