期刊
ANALYTICAL CHEMISTRY
卷 90, 期 20, 页码 12051-12058出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.analchem.8b02820
关键词
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资金
- National Key Research and Development Program of China [2016YFA0501602]
- NSFC [21775021, 21375015]
- Priority Discipline Development Program of Jiangsu Province
The combination of microbead array with assay chemistry of isothermal amplification enables the continuous development of nucleic acid detection techniques. Herein we report the implementation of ligation-rolling circle amplification (RCA) reaction on quantum dots-encoded microbead (Qbead) for the detection of multiplex G-6 quadruplex (G4) forming sequences. The reaction time of RCA on the Qbead was optimized to be 60 min. Zinc phthalocyanine (ZnPc), a molecular light switch, was selected as the G4-specific label. In the presence of target, the target-triggered ligation-RCA produced long DNA concatemer consisting of tandem repeats of G4-forming sequence, and the labeling helped generate G4/ZnPc nanowires on the Qbead. With the G4/ZnPc nanowires as fluorescent labels, the array of three encoded Qbeads was capable of detecting three G4-forming sequences by flow cytometry in a high throughput and specific manner. Alternatively, with the G4/ZnPc nanowires as catalytic labels, chemiluminescence of H2O2 mediated oxidation of luminol could be used for detecting the target G4-forming sequences with high sensitivity. The catalytic chemiluminescence achieved a limit of detection of 0.5 ng of genomic DNA with 5 logs of linear dynamic range for the detection of the blood sample of a myeloproliferative neoplasms patient. Together the proposed isothermal amplification-on-Qbead assay featured robust detection platform, significant signal amplification, and flexible detection strategy, holding high potential in application in large-scale or focused nucleic acid testing.
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