Enzyme-free and multiplexed microRNA detection using microRNA-initiated DNA molecular motor

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• 作者：Hui Wang ; Honghong Wang ; Chenghui Liu ; Xinrui Duan ; Zhengping Li
• 关键词：multiplexed microRNA detection ; DNA molecular motor ; enzyme ; free ; synchronous fluorescence spectrum
• 刊名：SCIENCE CHINA Chemistry
• 出版年：2016
• 出版时间：January 2016
• 年：2016
• 卷：59
• 期：1
• 页码：83-88
• 全文大小：844 KB
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• 作者单位：Hui Wang (1) (2) (3)
  Honghong Wang (1) (2) (3)
  Chenghui Liu (1) (2) (3)
  Xinrui Duan (1) (2) (3)
  Zhengping Li (1) (2) (3)
  
  1. Key Laboratory of Applied Surface and Colloid Chemistry, Ministry of Education, Xi’an, 710062, China
  2. Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province, Xi’an, 710062, China
  3. School of Chemistry and Chemical Engineering, Shaanxi Normal University, Xi’an, 710062, China
  
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Chemistry
  Chinese Library of Science
  Chemistry
  
• 出版者：Science China Press, co-published with Springer
• ISSN：1869-1870

文摘

In this work, we have developed a sensitive, simple, and enzyme-free assay for detection of microRNAs (miRNAs) by means of a DNA molecular motor consisting of two stem-loop DNAs with identical stems and complementary loop domains. In the presence of miRNA target, it can hybridize with one of the stem-loop DNA to open the stem and to produce a miRNA/DNA hybrid and a single strand (ss) DNA, the ssDNA will in turn hybridize with another stem-loop DNA and finally form a double strand (ds) DNA to release the miRNA. One of the stem-loop DNA is double-labeled by a fluorophore/quencher pair with efficiently quenched fluorescence. The formation of dsDNA can produced specific fluorescence signal for miRNA detection. The released miRNA will continuously initiate the next hybridization of the two stem-loop DNAs to form a cycle-running DNA molecular motor, which results in great fluorescence amplification. With the efficient signal amplification, as low as 1 pmol/L miRNA target can be detected and a wide dynamic range from 1 pmol/L to 2 nmol/L is also obtained. Moreover, by designing different stem-loop DNAs specific to different miRNA targets and labeling them with different fluorophores, multiplexed miRNAs can be simultaneously detected in one-tube reaction with the synchronous fluorescence spectrum (SFS) technique.