The Effect of Monovalent Cation Binding on DNA Hairpin Stability

在这项由分析和表面化学计划支持的研究中，Stellwagen教授和她的团队将利用毛细管电泳法研究单价阳离子与DNA发夹的结合，以及单价阳离子结合对发夹稳定性的影响。目前的结构预测算法不能可靠地预测DNA发夹的稳定性，这表明导致发夹形成的相互作用没有正确地包括在折叠程序中。由于单链DNA低聚体中的发夹结构只有微弱的稳定性，因此很难用传统的方法，如紫外线吸收或差示扫描量热法来研究它们。相比之下，自由溶液毛细管电泳能直接检测发夹的形成和阳离子结合，因为电泳迁移率取决于分析物的形状和有效电荷。拟议研究的更广泛影响：该项目的一个重要方面是对生物化学和(或)生物物理科学方面的本科生和应届毕业生的培训。还有更广泛的科学和技术影响。单链DNA寡聚体中意外的发夹形成会干扰DNA寡聚体与其目标DNA或RNA序列的杂交，给多重PCR反应的设计和微阵列实验的解释带来困难。发夹的形成也会干扰DNA寡核苷酸作为反义基因治疗剂的有效性。因此，能够预测DNA低聚体中发夹的形成并了解影响发夹稳定性的因素是很重要的。我们开发的用于测量一价阳离子与DNA结合的毛细管电泳法是通用的，可以用于测量其他配体与其他分析物的结合。。

In this research supported by the Analytical and Surface Chemistry Program, Professor Stellwagen and her group will study the binding of monovalent cations to DNA hairpins, and the effect of monovalent cation binding on hairpin stability, using capillary electrophoresis. The current structure-prediction algorithms do not reliably predict DNA hairpin stability, indicating that the interactions leading to hairpin formation are not properly included in the folding routines. Since hairpin structures in single-stranded DNA oligomers are only marginally stable, they are difficult to study by conventional methods such as ultraviolet absorption or differential scanning calorimetry. By contrast, free solution capillary electrophoresis can detect hairpin formation and cation binding directly, because electrophoretic mobility depends on both the shape and the effective charge of the analyte. Broader impact of the proposed research: An important aspect of the project is its contribution to the training of undergraduate students and recent graduates in the biochemical and/or biophysical sciences. There are also scientific and technological broader impacts. Unexpected hairpin formation in single-stranded DNA oligomers can interfere with the hybridization of DNA oligomers to their target DNA or RNA sequences, creating difficulties in the design of multiplex PCR reactions and the interpretation of microarray experiments. Hairpin formation can also interfere with the effectiveness of DNA oligonucleotides used as antisense gene therapy agents. Hence, it is important to be able to predict hairpin formation in DNA oligomers and to understand the factors contributing to hairpin stability. The capillary electrophoresis methods that we have developed to measure monovalent cation binding to DNA are of general utility and can be used to measure the binding of other ligands to other analytes. .