Competition dialysis: A method for the study of structural selective nucleic acid binding
Competition dialysis: A method for the study of structural selective nucleic acid binding
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DOI：
10.1016/j.ymeth.2006.09.010
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发表时间：
2007-06-01
期刊：
影响因子：
4.8
通讯作者：
Chaires, Jonathan B.
中科院分区：
文献类型：
--
作者：
Ragazzon, Patricia A.;Garbett, Nichola C.;Chaires, Jonathan B.
竞争透析是一个强大的新的发现配体结合的核酸与结构或序列的选择性。该方法是基于坚定的热力学原理，是简单的实现。在竞争透析实验中，将核酸结构和序列的阵列针对共同的测试配体溶液透析。平衡后，通过吸光度或荧光测量确定与每个结构或序列结合的配体的量。由于所有结构和序列与相同的游离配体浓度处于平衡状态，因此结合量与配体结合亲和力成正比。因此，竞争透析提供了选择性的直接和定量测量，并且明确地识别阵列内的哪些样品是特定配体优选的。我们在这里描述的第三代实现的方法，其中竞争透析适用于在96孔板格式。在这种形式下，我们已经能够大大扩展所研究的核酸结构阵列，现在可以常规地研究感兴趣的配体与46种不同结构和序列的相互作用。(c)2006年爱思唯尔公司All rights reserved.
Competition dialysis is a powerful new too] for the discovery of ligands that bind to nucleic acids with structural- or sequence-selectivity. The method is based on firm thermodynamic principles and is simple to implement. In the competition dialysis experiment, an array of nucleic acid structures and sequences is dialyzed against a common test ligand solution. After equilibration, the amount of ligand bound to each structure or sequence is determined by absorbance or fluorescence measurements. Since all structures and sequences are in equilibrium with the same free ligand concentration, the amount bound is directly proportional to the ligand binding affinity. Competition dialysis thus provides a direct and quantitative measure of selectivity, and unambiguously identifies which of the samples within the array are preferred by a particular ligand. We describe here the third generation implementation of the method, in which competition dialysis was adapted for use in a 96-well plate format. In this format, we have been able to greatly expand the array of nucleic acid structures studied, and now can routinely study the interactions of a ligand of interest with 46 different structures and sequences. (c) 2006 Elsevier Inc. All rights reserved.