Zheng Xiangjiang, Niu Li, Wei Di, Li Xuemei, Zhang Shusheng
Key Laboratory of Sensor Analysis of Tumor Marker, Ministry of Education, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, P. R. China.
Shandong Provincial Key Laboratory of Detection Technology for Tumor Makers, Research Institute of Biochemical Analysis, Linyi University, Linyi 276000, P. R. China.
Sci Rep. 2016 Oct 25;6:35982. doi: 10.1038/srep35982.
MicroRNA (miRNA) was a promising class of cancer biomarkers. Here we developed a label-free method for sensitive measurement of let-7d miRNA based on multiple amplification techniques. The primer will bind to the duplex strand DNA that was formed by stem-loop template and target let-7d to initiate strand displacement amplification (SDA) in tandem. The released single strand DNA will be a primer to bind the circular template to initiate rolling circle amplification (RCA). The products based on multiple amplifications will be detected by a standard fluorimeter with N-methyl mesoporphyrin IX (NMM) as the fluorescent indicator. The proposed method exhibited excellent selectivity and high sensitivity with a detection limit of as low as 1.5 × 10 M. Moreover, this methodology was used for the determination of biomolecules in real serum samples with satisfying results.
微小RNA（miRNA）是一类很有前景的癌症生物标志物。在此，我们基于多种扩增技术开发了一种用于灵敏检测let-7d miRNA的无标记方法。引物将与由茎环模板和靶标let-7d形成的双链DNA结合，以串联方式启动链置换扩增（SDA）。释放的单链DNA将作为引物与环状模板结合，启动滚环扩增（RCA）。基于多次扩增的产物将通过以N-甲基中卟啉IX（NMM）作为荧光指示剂的标准荧光计进行检测。所提出的方法表现出优异的选择性和高灵敏度，检测限低至1.5×10⁻⁹ M。此外，该方法用于实际血清样品中生物分子的测定，结果令人满意。
