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Microchimica Acta

, 186:531 | Cite as

Graphene oxide-based fluorometric determination of microRNA-141 using rolling circle amplification and exonuclease III-aided recycling amplification

  • Mei LiEmail author
  • Xiong Xu
  • QingYou Cai
  • XuJian Luo
  • ZhongGao Zhou
  • GuoHai Xu
  • YongRong XieEmail author
Original Paper
  • 43 Downloads

Abstract

A graphene oxide-based method has been developed for ultrasensitive and selective determination of microRNA-141 by means of rolling circle amplification (RCA) and exonuclease III (Exo III)-assisted recycling amplification. The method uses (a) a padlock probe with a hybrid sequence that is complementary to the target microRNA-141 at both the 5'- and the 3'-end, and (b) a long binding region of a signalling reporter strand. On addition of microRNA-141, it acts as the primer for triggering the RCA reaction following ligation. This results in the formation of a repeatedly concatenated sequence of the padlock probe. Subsequently, the RCA product hybridizes with the fluorescein-labelled signal strand to form the duplex DNA containing the blunt 3'-termini of signal strand. Addition of Exo III causes signal strand digestion and leads to RCA product recycling and liberation of fluorescein. Added graphene oxide does not absorb the fluorescein liberated because of the poor mutual interaction. Therefore, microRNA-141 can be quantified by measurement of the green fluorescence under excitation/emission wavelengths of 470/520 nm. The method has a 100 aM detection limit towards microRNA-141 and works in the wide range from 1 fM to 1 nM. It can discriminate even single-mismatched microRNA and shows good selectivity and sensitivity when applied to spiked human serum.

Graphical abstract

Schematic representation of a graphene oxide (GO)-based method for fluorometric determination of microRNA by using rolling circle amplification and exonuclease III (Exo III)-aided recycling amplification. With microRNA, the fluorescein-labelled signal strand becomes digested, and this genererates a fluorescent signal.

Keywords

MicroRNA Signal amplification DNA polymerase Exo III digestion Graphene oxide quenching Padlock probe Serum samples Fluorescein-label Signal strand Fluorescence detection 

Notes

Acknowledgements

This work was supported by the Natural Science Foundation of China (21461002, 21241005 and 21562002), Key Laboratory of Jiangxi University for Functional Materials Chemistry (FMC15104), Jiangxi Provincial Department of Education (GJJ170826 and JXJG-16-14-6), and Gannan Normal University (16zb19).

Compliance with ethical standards

The author(s) declare that they have no competing interests.

Supplementary material

604_2019_3676_MOESM1_ESM.doc (588 kb)
ESM 1 (DOC 588 kb)

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Copyright information

© Springer-Verlag GmbH Austria, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Key Laboratory of Jiangxi University for Functional Material Chemistry, College of Chemistry & Chemical EngineeringGannan Normal UniversityGanzhouChina
  2. 2.College of Mathematics and Computer SciencesGannan Normal UniversityGanzhouChina
  3. 3.School of Chemical & Food ScienceYulin Normal UniversityYulinChina

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