Graphene oxide and enzyme-assisted dual-cycling amplification method for sensitive fluorometric determination of DNA

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.

Schematic representation of a GO-based, Exo III-assisted method for dual-signal amplified detection of DNA, for which human haemochromatosis protein (HFE) gene is designed as the model target. The assay involves graphene oxide (GO), exonuclease (Exo III), and two specially designed, fluorophore-labelled hairpin probes (HP1 and HP2).

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Acknowledgements

This work was partially supported by the Research Grants Council of the Hong Kong Special Administrative Region under Grant No. 16228216 and the National Natural Science Foundation of China under Grant No. 21705076. We would also like to acknowledge the supply of graphene oxide from World Linkage Holdings Limited (Hong Kong).

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Correspondence to Zhigang Li or Jiahao Huang.

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Iwe, I., Li, Z. & Huang, J. Graphene oxide and enzyme-assisted dual-cycling amplification method for sensitive fluorometric determination of DNA. Microchim Acta 186, 716 (2019). https://doi.org/10.1007/s00604-019-3825-7

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Keywords

  • DNA sensor
  • Fluorescence
  • Exonuclease III
  • Nanomaterial
  • Human hemochromatosis protein gene
  • Fetal bovine serum