Graphene oxide and enzyme-assisted dual-cycling amplification method for sensitive fluorometric determination of DNA
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ORIGINAL PAPER
Graphene oxide and enzyme-assisted dual-cycling amplification method for sensitive fluorometric determination of DNA Idorenyin Iwe 1 & Zhigang Li 1 & Jiahao Huang 2 Received: 3 May 2019 / Accepted: 14 September 2019 # Springer-Verlag GmbH Austria, part of Springer Nature 2019
Abstract A fluorometric method is described for the determination of DNA. It involves the use of graphene oxide (GO), exonuclease III (Exo III), and two specially designed fluorophore-labeled hairpin probes (HP1 and HP2). Different from other GO-based DNA assays, the method takes advantage of the distinct binding abilities of GO with hairpin DNA probes and single nucleotides. GO serves as a strong quencher for fluorescent labels to ensure a very low background signal. Two reaction cycles mediated by Exo III are employed to enhance the signals. The combination of GO-induced quenching and Exo III-mediated dual regeneration of analytes leads to a detection limit as low as 1 pM for the model analyte human hemochromatosis protein (HFE) gene. The method is also applicable for the determination of HFE gene spiked into fetal bovine serum. Keywords DNA sensor . Fluorescence . Exonuclease III . Nanomaterial . Human hemochromatosis protein gene . Fetal bovine serum
Introduction Sensitivity is a crucial issue in DNA sensing as the DNA concentration in real biosamples is usually too low to be easily quantified [1]. Therefore, it is highly desirable to develop methods with amplification principles that are sufficiently sensitive for DNA detection [2, 3]. Enzyme-free and enzyme-assisted methods have been widely employed for the amplified detection of DNA targets. Among them, catalytic hairpin assembly (CHA) [4, 5] and hybridization
Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00604-019-3825-7) contains supplementary material, which is available to authorized users. * Zhigang Li [email protected] * Jiahao Huang [email protected] 1
Department of Mechanical and Aerospace Engineering, The Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China
2
Guangdong Provincial Key Laboratory of Construction and Detection in Tissue Engineering, Biomaterials Research Center, School of Biomedical Engineering, Southern Medical University, Guangzhou 510515, China
chain reaction (HCR) [6] are two powerful enzyme-free strategies. In enzyme-aided systems, popular amplification technologies include polymerase chain reaction (PCR) [7], ligase chain reaction (LCR) [8], rolling circle amplification (RCA) [9], and loop-mediated isothermal amplification (LAMP) [10]. To further improve the detection sensitivity, nanomaterials with extraordinary properties are also promising choices, which have been incorporated into various sensing platforms [11]. Popular nanomaterials include gold nanoparticles (AuNPs) [12], quantum dots (QDs) [13], carbon nanotubes (CNTs) [14], gold nanowires [9], molybdenum disulphide (MoS2) [3], and graphene oxide (GO) [15, 16]. Special attention has been paid
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