Intra-molecular G-quadruplex structure generated by DNA-templated click chemistry: “Turn-on” fluorescent probe for copper ions

• Published in: Biosensors & bioelectronics 2014-05, Vol.55, p.187-194
• Main Authors: Shen, Qinpeng, Zhou, Lifen, Yuan, Yijia, Huang, Yan, Xiang, Binbin, Chen, Chunyan, Nie, Zhou, Yao, Shouzhuo
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biotechnology; Click chemistry; Click Chemistry - methods; Copper - analysis; Copper - chemistry; Copper ion; Crystal violet (CV); DNA - chemistry; DNA - ultrastructure; DNA-templated organic synthesis (DTS); Fluorescent Dyes - analysis; Fluorescent Dyes - chemical synthesis; Fundamental and applied biological sciences. Psychology; G-quadruplex; G-Quadruplexes; Ions; Molecular Imprinting - methods; Reproducibility of Results; Sensitivity and Specificity; Spectrometry, Fluorescence - methods
• ISSN: 0956-5663; 1873-4235
• DOI: 10.1016/j.bios.2013.12.019

A novel homogenous fluorescent sensor for signal-on detection of Cu2+ has been developed based on intra-molecular G-quadruplex formed by DNA-templated click reaction and crystal violet (CV) as label-free signal reporter. The clickable DNA probe consists of two G-rich strands (A and B) bearing azide and alkyne group, respectively, and a template strand (C) locating two proximate reactants by pairing with A and B. The sequences of A and B are derived from asymmetric split of the G-quadruplex sequence (TTAGGG)4. In the presence of Cu2+, the whole G-quadruplex sequence A–B is generated by chemical ligation of A and B via copper ion-catalyzed alkyne-azide cycloaddition, then released from template by toehold strand displacement, and consequently forming a stable intra-molecular G-quadruplex, which binds with CV to generate a strong fluorescent signal. Oppositely, weak fluorescence was obtained without Cu2+ because of unstable intermolecular G-quadruplex formed by A and B and lack of lateral loop connection. Therefore, the Cu2+ can be sensitively and specifically detected by the fluorescence of the CV-stained G-quadruplex with a low detection limit of 65nM and a linear range of 0.1–3µM. This method rationally integrated the DNA-templated synthesis and G-quadruplex structure-switch, presenting a simple and promising approach for biosensor development. •A novel fluorescent copper ion biosensor relied on DNA-templated click reaction mediated intra-molecular G-quadruplex structure formation.•This “mix-and-read” signal-on detection is straightforward and homogenous based on the G-quadruplex-specific fluorogenic property of crystal violet.•High sensitivity due to the high yielding capability of DNA-templated organic synthesis (DTS).•Highly selective for copper ion because of the great specificity of click chemistry.