College of Chemistry, Chemical Engineering and Materials Science, Key Laboratory of Molecular and Nano Probes, Ministry of Education, Shandong Normal University, 88 Wenhua East Road, Jinan 250014, China.
Biosens Bioelectron. 2013 Mar 15;41:907-10. doi: 10.1016/j.bios.2012.10.007. Epub 2012 Oct 8.
A bifunctional fluorescent oligonucleotide probe for small molecules and protein detection has been developed based on turn on fluorescence response via the target induced structure-switching of molecular beacon. The two loops of this molecular beacon are designed in such a manner that they consist of thrombin (Tmb) aptamer sequence and adenosine triphosphate (ATP) aptamer sequence, respectively, which are utilized to sense thrombin and ATP. The oligonucleotide forms double stem-loops in the absence of targets, yielding no fluorescence emission because of the FRET from the excited fluorophore to the proximal quencher. Upon addition of the target, the ATP or Tmb, its specific interaction with loop sequence of the hairpin structure induce the separation of reporter fluorophore and the fluorescence quencher of the molecular beacon, resulting in an increase of fluorescence response. Hence, the separate analysis of ATP and Tmb could be realized through only one designed molecular beacon. The detection limits were estimated to be 25 nM for ATP and 12 nM for Tmb, respectively. The results of this study should substantially broaden the perspective for future development of oligonucleotide probe for analysis of other analytes.
基于分子信标通过靶标诱导的结构切换实现的荧光开启响应，开发了一种用于小分子和蛋白质检测的双功能荧光寡核苷酸探针。该分子信标的两个环分别设计为包含凝血酶（Tmb）适体序列和三磷酸腺苷（ATP）适体序列，分别用于检测凝血酶和 ATP。在没有靶标的情况下，寡核苷酸形成双茎环结构，由于从激发荧光团到近端猝灭剂的 FRET，没有荧光发射。当加入靶标（ATP 或 Tmb）时，其与发夹结构环序列的特异性相互作用导致报告荧光团和分子信标的荧光猝灭剂分离，从而导致荧光响应增加。因此，通过仅一个设计的分子信标即可实现对 ATP 和 Tmb 的单独分析。ATP 的检测限估计为 25 nM，Tmb 的检测限估计为 12 nM。这项研究的结果应该大大拓宽未来用于分析其他分析物的寡核苷酸探针的发展前景。
