Monroy-Contreras Ricardo, Vaca Luis
Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, 04510 Mexico, DF, Mexico.
J Nucleic Acids. 2011;2011:741723. doi: 10.4061/2011/741723. Epub 2011 Aug 22.
Recent advances in RNA functional studies highlights the pivotal role of these molecules in cell physiology. Diverse methods have been implemented to measure the expression levels of various RNA species, using either purified RNA or fixed cells. Despite the fact that fixed cells offer the possibility to observe the spatial distribution of RNA, assays with capability to real-time monitoring RNA transport into living cells are needed to further understand the role of RNA dynamics in cellular functions. Molecular beacons (MBs) are stem-loop hairpin-structured oligonucleotides equipped with a fluorescence quencher at one end and a fluorescent dye (also called reporter or fluorophore) at the opposite end. This structure permits that MB in the absence of their target complementary sequence do not fluoresce. Upon binding to targets, MBs emit fluorescence, due to the spatial separation of the quencher and the reporter. Molecular beacons are promising probes for the development of RNA imaging techniques; nevertheless much work remains to be done in order to obtain a robust technology for imaging various RNA molecules together in real time and in living cells. The present work concentrates on the different requirements needed to use successfully MB for cellular studies, summarizing recent advances in this area.
RNA功能研究的最新进展凸显了这些分子在细胞生理学中的关键作用。人们已经采用了多种方法来测量各种RNA种类的表达水平，这些方法使用的是纯化的RNA或固定细胞。尽管固定细胞能够观察RNA的空间分布，但为了进一步了解RNA动态在细胞功能中的作用，仍需要具备实时监测RNA转运到活细胞能力的检测方法。分子信标（MBs）是具有茎环发夹结构的寡核苷酸，一端配备有荧光淬灭剂，另一端配备有荧光染料（也称为报告基团或荧光团）。这种结构使得在没有其靶标互补序列的情况下，MB不发出荧光。与靶标结合后，由于淬灭剂和报告基团的空间分离，MB会发出荧光。分子信标是用于开发RNA成像技术的有前景的探针；然而，为了获得一种能够在活细胞中实时同时成像各种RNA分子的强大技术，仍有许多工作要做。目前的工作集中在成功使用MB进行细胞研究所需的不同要求上，总结了该领域的最新进展。
