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Please use this identifier to cite or link to this item: http://ntour.ntou.edu.tw:8080/ir/handle/987654321/38481

Title: Detection of microRNA in Tumor Cells using Exonuclease III and Graphene Oxide-Regulated Signal Amplification
Authors: Rong-Cing Huang
Wei-Jane Chiu
Yu-Jia Li
Chih-Ching Huang
Contributors: 國立臺灣海洋大學:生命科學暨生物科技學系
Keywords: tumor cells
graphene oxide
mass spectrometry
microRNA
oligonucleotides
single-nucleotide polymorphism
Date: 2014
Issue Date: 2016-09-07T06:39:13Z
Publisher: ACS Applied Materials and Interfaces
Abstract: Abstract: In this study, we developed a label-free, ultrasensitive graphene oxide (GO)-based probe for the detection of oligonucleotides by laser desorption/ionization mass spectrometry (LDI-MS). On the basis of simple π–π stacking and electrostatic interactions between rhodamine 6G (R6G) and GO, we prepared the nanocomposite R6G-modified GO (R6G-GO). Signal intensities of R6G increased in mass spectra in the presence of single-stranded oligonucleotides under pulsed laser irradiation (355 nm) of R6G-GO. In addition, the signal intensity of R6G was stronger in the presence of short oligonucleotides. Because small oligonucleotides improve the LDI efficiency of R6G on GO, we designed an enzyme-amplified signal transduction probe system for the detection of microRNA (miRNA). After specific digestion of the probe DNA (pDNA) strand from pDNA/miRNA-hybridized complexes by exonuclease III (Exo III), the resulting small oligonucleotide fragments increased the R6G signal during LDI-MS of R6G-GO. In addition, the signal intensity of the R6G ions increased with increasing concentrations of the target miRNA. Coupling this enzyme reaction and R6G-GO with LDI-MS enabled the detection of miRNA at concentrations of the femtomolar (fM) level. We also demonstrated the analysis of miRNA in tumor cells and utilized this R6G-GO probe in the detection of a single-nucleotide polymorphism (SNP) in the Arg249Ser unit of the TP53 gene. This simple, rapid, and sensitive detection system based on the coupling of functional GO with LDI-MS appears to have great potential as a tool for the bioanalyses of oligonucleotides and proteins.
Relation: 6(24)
URI: http://ntour.ntou.edu.tw:8080/ir/handle/987654321/38481
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