Superior structure stability and selectivity of hairpin nucleic acid probes with an L-DNA stem.
Superior structure stability and selectivity of hairpin nucleic acid probes with an L-DNA stem.
复制标题
具有L-DNA茎的发夹核酸探针的优质结构稳定性和选择性。
DOI：
10.1093/nar/gkm771
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发表时间：
2007
影响因子：
14.9
通讯作者：
Tan, Weihong
中科院分区：
文献类型：
--
作者：
Kim, Youngmi;Yang, Chaoyong James;Tan, Weihong
发夹核酸探针在许多领域中非常有用，特别是用于细胞内和体外核酸检测。这些探针的成功可归因于它们在靶结合时的构象变化可与多种信号转导机制偶联的容易性。然而，假阳性信号主要由于热波动和茎侵入而从发夹的打开中产生。当茎与其互补序列相互作用时，茎侵入发生，并且在复杂的生物样品中特别成问题。为了解决发夹探针中茎侵入的问题，我们已经创建了一个修饰的分子信标，该分子信标在茎中掺入非天然对映体l-DNA，在环中掺入天然d-DNA或2′-O-Me修饰的RNA。除了l-DNA不能与d-DNA形成稳定的双链体之外，l-DNA具有与d-DNA相同的物理特性。在这里，我们表明，将L-DNA纳入干区域的分子信标减少内和分子间干入侵，增加熔解温度，提高其目标的选择性，并导致增强的生物稳定性。我们的研究结果表明，l-DNA是有用的设计功能的核酸探针，特别是生物应用。
Hairpin nucleic acid probes have been highly useful in many areas, especially for intracellular and in vitro nucleic acid detection. The success of these probes can be attributed to the ease with which their conformational change upon target binding can be coupled to a variety of signal transduction mechanisms. However, false-positive signals arise from the opening of the hairpin due mainly to thermal fluctuations and stem invasions. Stem invasions occur when the stem interacts with its complementary sequence and are especially problematic in complex biological samples. To address the problem of stem invasions in hairpin probes, we have created a modified molecular beacon that incorporates unnatural enantiomeric l-DNA in the stem and natural d-DNA or 2′-O-Me-modified RNA in the loop. l-DNA has the same physical characteristics as d-DNA except that l-DNA cannot form stable duplexes with d-DNA. Here we show that incorporating l-DNA into the stem region of a molecular beacon reduces intra- and intermolecular stem invasions, increases the melting temperature, improves selectivity to its target, and leads to enhanced bio-stability. Our results suggest that l-DNA is useful for designing functional nucleic acid probes especially for biological applications.