Key Laboratory of Chemical Sensing & Analysis in Universities of Shandong, School of Chemistry and Chemical Engineering, University of Jinan, Jinan, Shandong 250022, China.
School of Biological Science and Technology, University of Jinan, Jinan, Shandong 250022, China.
Biosens Bioelectron. 2017 Mar 15;89(Pt 2):710-714. doi: 10.1016/j.bios.2016.10.099. Epub 2016 Nov 11.
MicroRNAs (miRNAs) play significant roles in a diverse range of biological progress and have been regarded as biomarkers and therapeutic targets in cancer treatment. Here, we develop a rapid and sensitive miRNA assay on the basis of exponential isothermal amplification in combination with the hairpin probes, which was designed for sensitive detection of miRNA. The binding of target miRNA with a linear DNA template initiates exponential isothermal amplification reaction (EXPAR) and generates the universal triggers which are complementary to the 3' protruding end of hairpin probe1(HP1). These triggers function not only as the primers to unfold the hairpin probes through catalysed hairpin assembly(CHA), generating distinct fluorescence signals, but also as the primer to initiate the next EXPAR. Moreover, CHA can release new triggers to initiate EXPAR or CHA. Thus this hairpin probes-mediated exponential isothermal amplification assay exhibits high sensitivity with a detection limit of 3.0×10M. More importantly, the isothermal condition and simple fluorescence measurement would greatly promote the development of a fast, point-of-care detection system. It can be completed in an hour which can effectively avoid miRNA from degradation. This hairpin probe-based circular exponential amplification assay holds a great promise for further application in biomedical research and early clinical diagnosis.
微小 RNA(miRNA)在多种生物过程中发挥着重要作用,被认为是癌症治疗中的生物标志物和治疗靶点。在这里,我们基于指数等温扩增结合发夹探针开发了一种快速灵敏的 miRNA 检测方法,用于灵敏检测 miRNA。靶 miRNA 与线性 DNA 模板的结合启动指数等温扩增反应(EXPAR),并产生与发夹探针 1(HP1)3'突出末端互补的通用触发物。这些触发物不仅可以作为通过催化发夹组装(CHA)展开发夹探针的引物,产生独特的荧光信号,还可以作为启动下一个 EXPAR 的引物。此外,CHA 可以释放新的触发物来启动 EXPAR 或 CHA。因此,这种发夹探针介导的指数等温扩增检测方法具有很高的灵敏度,检测限为 3.0×10M。更重要的是,等温条件和简单的荧光测量将极大地促进快速、即时检测系统的发展。它可以在一小时内完成,这可以有效地避免 miRNA 的降解。这种基于发夹探针的循环指数扩增检测方法在生物医学研究和早期临床诊断中有很好的应用前景。
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