A simple enzyme-assisted cascade amplification strategy for ultrasensitive and label-free detection of DNA

作者：Wang, Kun; Zhai, Fu-Heng; He, Meng-Qi; Wang, Jin; Yu, Yong-Liang^*; He, Rong-Huan^*

来源：Analytical and Bioanalytical Chemistry, 2019, 411(19): 4569-4576.

DOI：10.1007/s00216-018-1422-y

• Biosensor
• Cascade
• Label-free
• Hairpin DNA
• Exonuclease III

A simple fluorescence biosensor is developed based on the enzyme-assisted cascade amplification strategy. The amplification system consists of a hairpin-structure DNA (H-DNA) and exonuclease III. The target DNA can hybridize with the H-DNA and initiate exonuclease III-assisted target recycling amplification to generate abundant G-rich DNA (G-DNA). One region of G-DNA is designed to possess the same sequence as target DNA. Thus, the G-DNA can also hybridize with H-DNA and initiate the digestion of H-DNA. The cascade strategy in this amplification system causes the concentration of G-DNA to grow exponentially. The fluorescence intensity of N-methylmesoporphyrin IX (NMM) is highly enhanced due to the formation of G-quadruplex configuration. Under optimal conditions, the cascade system could achieve an admirable sensitivity with a detection limit of 52fM for HIV DNA, and guarantees a satisfactory specificity. Moreover, the cascade system could be implemented for other target DNA detections by substituting the recognition region of the H-DNA. In this way, a detection limit of 65fM for HBV DNA could be achieved by the cascade system. The target DNA analysis in a real serum sample further indicates that this biosensor has potential for future application in clinical diagnosis.

• 出版日期2019-7
• 单位东北大学