Modulating fluorescence anisotropy of dye-labeled DNA without involving mass amplification
Modulating fluorescence anisotropy of dye-labeled DNA without involving mass amplification
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调节染料标记 DNA 的荧光各向异性，无需进行质量扩增
DOI：
10.1016/j.talanta.2016.01.016
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发表时间：
2016-07-01
期刊：
影响因子：
6.1
通讯作者：
Li, Na
中科院分区：
文献类型：
--
作者：
Pei, Xiaojing;Huang, Hongduan;Li, Na
荧光各向异性是一种简单、均匀和经济的分析技术，是研究与分子相互作用相关的染料微环境变化的宝贵技术。深入了解影响荧光各向异性信号的变量有助于更好的实验设计，从而有效地提高分析性能。这项工作是评估可能显著影响荧光各向异性的因素的后续工作。基于染料与DNA的相互作用，系统地研究了末端标记、中位标记和多种标记方式下双链DNA荧光各向异性随长度变化的规律。通过标记dsDNA中间的荧光团和增加dsDNA上的标记数，可以扩大荧光各向异性值和碱基对响应的动态范围。末端标记方式的C悬臂构型可以增强荧光各向异性信号，但不能扩大碱基对的响应范围。所有标记荧光团的结果都加强了平坦效应，即当DNA双链的长度大于临界碱基对数量时，荧光各向异性信号对DNA双链长度的增加没有响应。这些发现为在各种应用中为简单和通用的荧光各向异性设计选择合适的荧光染料和标记位置提供了视角。(C)2016爱思唯尔B.V.保留所有权利。
Fluorescence anisotropy, known as a simple, homogeneous and cost-effective analytical technology, is an invaluable technique for studying the micro -environmental changes of the dye associated with the molecular interactions. An in-depth understanding of the variables affecting the fluorescence anisotropy signal can facilitate better experimental designs to effectively improve the analytical performance. This work is a follow-up effort in evaluating the factors that can significantly influence fluorescence anisotropy. We systematically studied fluorescence anisotropy of dsDNA with the changing length based on dye-DNA interactions, with the fluorophores in the end -labeling, the middle -site -labeling, and multiple number of labeling manners. The fluorescence anisotropy value and the base -pair response dynamic range could be expanded by labeling the fluorophores in the middle of dsDNA and increasing the number of labels on dsDNA. The C overhang configuration in the end -labeling manner could enhance the fluorescence anisotropy signal but not expand the base -pair response range. Results from all the labeling fluorophores reinforced the leveling -off effect, i.e., the fluorescence anisotropy signal does not response to the increased length of the DNA duplex when the length is larger than a critical number of base pairs. These findings provide perspectives about choosing appropriate fluorescent dyes and labeling sites for simple and universal fluorescence anisotropy designs in various applications. (C) 2016 Elsevier B.V. All rights reserved.