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Ratiometric fluorescent 3D DNA walker and catalyzed hairpin assembly for determination of microRNA

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Abstract

Using 6-carboxyfluorescein (FAM) and tetramethyl rhodamine (TAMRA) as fluorescent signals a ratiometric fluorescent three-dimensional (3D) DNA walker based on a catalytic hairpin assembly (CHA) reaction for microRNA-122 detection was constructed. This method uses CHA reaction triggered indirectly by the target to mediate the 3D DNA walker operation to amplify the signal. The dual emission ratio fluorescent signal with a single excitation wavelength was used as the signal output. This strategy combines DNA walker with CHA reaction and proportional fluorescence signal output methods, which can effectively reduce the background fluorescence signal and the risk of generating false-positive signals. Thus, the impact of environmental factors on the experiment is reduced, thereby obtaining reliable and stable experimental results. It uses the fluorescence excitation wavelength of 488 nm and the maximum fluorescence emission wavelength of 520 nm and 580 nm, respectively. It has a good linear response at a microRNA concentration range of 156.0 pM ~ 7.00 nM and a detection limit of 42.94 pM. This strategy has been successfully applied to detect microRNAs in spiked serum samples.

Schematic representation of three-dimensional (3D) DNA walker constructed using catalytic hairpin self-assembly reaction (CHA)-assisted amplification and ratiometric fluorescence signal output for the detection of miRNA-122 closely related to hepatitis

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Funding

This work was supported by the National Natural Science Foundation of China (no. 21465007, 21565007, 21165004, 21163002), the Guangxi Natural Science Foundation of China (2015GXNSFGA139003), the Bagui group of Guangxi province, and the project of State Key Laboratory for Chemistry and Molecular Engineering of Medicinal Resources, School of Chemistry and Pharmaceutical Science of Guangxi Normal University (CMEMR2014-A08).

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Authors and Affiliations

  1. Key Laboratory for the Chemistry and Molecular Engineering of Medicinal Resources (Ministry of Education of China), School of Chemistry and Pharmaceutical Science of Guangxi Normal University, Guilin, 541004, China

    Qing Li, Xuehua Liang, Xiaomei Mu, Li Tan, Jiangnan Lu, Kun Hu, Shulin Zhao & Jianniao Tian

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  1. Qing Li
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Correspondence to Jianniao Tian.

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All procedures and experiments about human blood samples were performed in accordance with the Guidelines for Care and Use of Laboratory biological of Guangxi Normal University (Guilin, China) and approved by the biological Ethics Committee of China.

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Li, Q., Liang, X., Mu, X. et al. Ratiometric fluorescent 3D DNA walker and catalyzed hairpin assembly for determination of microRNA. Microchim Acta 187, 365 (2020). https://doi.org/10.1007/s00604-020-04324-5

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