Effects of Vicinal Surface Chemistry on DNA Base-Pairing using Single-Molecule RE

使用单分子 RE 邻位表面化学对 DNA 碱基配对的影响

基本信息

批准号：
8280932
负责人：
DANIEL SCHWARTZ
金额：
$ 17.59万
依托单位：
UNIVERSITY OF COLORADO
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-04-01 至 2014-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8280932
关键词：
Adsorption Affect Base Pairing Behavior Characteristics Chemicals Chemistry Complex Contrast Media DNA DNA Sequence Development Diagnostic Diffusion Electrostatics Energy Transfer Environment Equilibrium Event Exhibits Fluorescence Microscopy Gene Delivery Glass Hydrocarbons Hydrogen Bonding Hydrophobic Interactions Image Lead Methods Microscopy Modeling Modification Molecular Molecular Conformation Nanotechnology Nanotubes Nucleic Acids Oligonucleotides Phase Polyethylene Glycols Polymers Population Preparation Process Property Quantum Dots Resolution Silicon Solid Solutions Structure Surface Techniques Technology Time Work aqueous base capsule design improved interfacial member molecular assembly/self assembly molecular dynamics molecular recognition nanoparticle novel particle research study residence self assembly single molecule stem

项目摘要

DESCRIPTION (provided by applicant): The overall objective of this work is to develop methods that can provide mechanistic information about the ways in which the chemical functionality of a nearby surface affects nucleic acid base- pairing and hybridization, and to use these methods to understand how surface chemistry can be used to control DNA self-assembly. DNA-based biomedical nanotechnologies rely directly on Watson-Crick base-pairing to achieve molecular recognition capable of directing self-assembly for a variety of diagnostic and targeting applications. While base-pairing in the solution phase is relatively well-understood, in many DNA nanotechnologies, base-pairing/hybridization is intended to occur in the vicinity of one or more interfaces/surfaces that may promote competitive non- specific interactions with nucleic acids due to the particular vicinal surface chemistry. A better understanding of surface effects on hybridization will ultimately lead to improvements in a wide range of DNA-based technologies. In particular, it will enable the design and preparation of improved surface-modification strategies. We propose to develop advanced single-molecule tracking methods using total internal reflection fluorescence microscopy (TIRFM) with single-molecule resolution, where resonance energy transfer (RET) is used to obtain simultaneous conformational information. This approach can identify direct molecule-by-molecule correlations between conformation and dynamic interfacial events (e.g. adsorption/desorption, interfacial mobility, conformational fluctuations, hybridization), providing mechanistic understanding of how vicinal surface chemistry affects both specific and non-specific DNA interactions. These methods will be used to study DNA dynamics and base- pairing on model surfaces that represent the most important examples of competing non-covalent interactions. This two-year plan focuses on understanding the interfacial dynamic behavior of (1) oligonucleotides that can self-hybridize to form stem-loop secondary structures, and (2) complementary oligonucleotide pairs. PUBLIC HEALTH RELEVANCE: DNA hybridization, the specific recognition of a DNA sequence by its complementary sequence, is the fundamental enabling phenomenon underlying a wide range of biomedical nanotechnologies, including molecular diagnostics, gene delivery, DNA sequencing, and many others. In these technologies, DNA molecules are required to exhibit specific recognition in the presence of foreign surfaces and materials that can interfere due to competitive chemical interactions. This work involves the development of novel single-molecule microscopy methods that will lead to a detailed understanding of these competitive interactions, permitting the design of improved materials and technologies.

描述（由申请人提供）：这项工作的总体目标是开发能够提供有关附近表面的化学功能如何影响核酸碱基配对和杂交的机制信息的方法，并使用这些方法来理解如何利用表面化学来控制 DNA 自组装。基于 DNA 的生物医学纳米技术直接依赖 Watson-Crick 碱基配对来实现分子识别，从而能够指导自组装，用于各种诊断和靶向应用。虽然溶液相中的碱基配对相对容易理解，但在许多 DNA 纳米技术中，碱基配对/杂交旨在发生在一个或多个界面/表面附近，这可能促进与核酸的竞争性非特异性相互作用，这是由于特定的邻近表面化学。更好地了解表面对杂交的影响最终将导致各种基于 DNA 的技术的改进。特别是，它将能够设计和准备改进的表面改性策略。我们建议使用具有单分子分辨率的全内反射荧光显微镜（TIRFM）开发先进的单分子跟踪方法，其中使用共振能量转移（RET）来获得同时构象信息。这种方法可以识别构象和动态界面事件（例如吸附/解吸、界面迁移性、构象波动、杂交）之间的直接分子间相关性，从而提供对邻近表面化学如何影响特异性和非特异性 DNA 相互作用的机制理解。这些方法将用于研究模型表面上的 DNA 动力学和碱基配对，这些模型表面代表了竞争性非共价相互作用的最重要示例。这个为期两年的计划重点是了解（1）可以自杂交形成茎环二级结构的寡核苷酸和（2）互补寡核苷酸对的界面动态行为。公共卫生相关性：DNA 杂交是通过互补序列对 DNA 序列进行特异性识别，是多种生物医学纳米技术的基本实现现象，包括分子诊断、基因传递、DNA 测序等。在这些技术中，DNA 分子需要在存在可能因竞争性化学相互作用而干扰的异物表面和材料的情况下表现出特异性识别能力。这项工作涉及新型单分子显微镜方法的开发，这将有助于详细了解这些竞争性相互作用，从而可以设计改进的材料和技术。