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Sensors 2019, 19(6), 1298; https://doi.org/10.3390/s19061298

Visual Detection of Cucumber Green Mottle Mosaic Virus Based on Terminal Deoxynucleotidyl Transferase Coupled with DNAzymes Amplification

Shandong Provincial Key Laboratory of Detection Technology for Tumor Markers, School of Chemistry and Chemical Engineering, School of Life Science, Linyi University, Linyi 276005, China
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Received: 24 January 2019 / Revised: 3 March 2019 / Accepted: 12 March 2019 / Published: 14 March 2019
(This article belongs to the Special Issue Sensors for Toxins and Pathogen Detection 2019)
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Abstract

A simple, rapid, and sensitive visual detection method for observing cucumber green mottle mosaic virus was reported based on the template-independent polymerization activity of terminal deoxynucleotidyl transferase (TdT), coupled with the cascade amplification of Mg2+-dependent DNAzyme and hemin/G-quadruplex DNAzyme. Briefly, the hybridized dsDNA of T1/P1 was cut into two parts at its position of 5′-AA↓CG↑TT-3′ by the restricted enzyme AcII. The longer, newborn fragment originating from P1 was tailed at its 3’-end by oligo dG, and an intact enzymatic sequence of Mg2+-dependent DNAzyme was generated. The substrate sequence in the loop segment of the hairpin probe (HP) hybridized with the newborn enzymatic sequence and was cleaved into two parts in the presence of Mg2+. The locked G-quadruplex sequence in the stem segment of the HP was released, which catalyzed the oxidation of ABTS2- in the presence of H2O2, and the resulting solution turned green. A correlation between the absorbance and concentration of T1 was obtained in a range from 0.1 pM to 2 nM, with a detection limit of 0.1 pM. In addition to promoting a lower detection limit and shorter monitoring time, this method also demonstrated an excellent selectivity to single or double nucleotide changes. Therefore, the designed strategy provided a rapid and efficient platform for viral inspection and plant protection. View Full-Text
Keywords: cucumber green mottle mosaic virus; visual detection; DNAzyme; terminal deoxynucleotidyl transferase; cascade amplification cucumber green mottle mosaic virus; visual detection; DNAzyme; terminal deoxynucleotidyl transferase; cascade amplification
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Wang, Y.; Liu, J.; Zhou, H. Visual Detection of Cucumber Green Mottle Mosaic Virus Based on Terminal Deoxynucleotidyl Transferase Coupled with DNAzymes Amplification. Sensors 2019, 19, 1298.

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