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SARS-CoV-2 RNA detection with duplex-specific nuclease signal amplification
Liu,Meiqing1; Li,Haoran1,2; Jia,Yanwei1,2,3; Mak,Pui In1,2; Martins,Rui P.1,2,4
2021-02-01
Source PublicationMicromachines
ISSN2072-666X
Volume12Issue:2
Abstract

The emergence of the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a zoonotic pathogen, has led to the outbreak of coronavirus disease 2019 (COVID-19) pandemic and brought serious threats to public health worldwide. The gold standard method for SARSCoV- 2 detection requires both reverse transcription (RT) of the virus RNA to cDNA and then polymerase chain reaction (PCR) for the cDNA amplification, which involves multiple enzymes, multiple reactions and a complicated assay optimization process. Here, we developed a duplexspecific nuclease (DSN)-based signal amplification method for SARS-CoV-2 detection directly from the virus RNA utilizing two specific DNA probes. These specific DNA probes can hybridize to the target RNA at different locations in the nucleocapsid protein gene (N gene) of SARS-CoV-2 to form a DNA/RNA heteroduplex. DSN cleaves the DNA probe to release fluorescence, while leaving the RNA strand intact to be bound to another available probe molecule for further cleavage and fluorescent signal amplification. The optimized DSN amount, incubation temperature and incubation time were investigated in this work. Proof-of-principle SARS-CoV-2 detection was demonstrated with a detection sensitivity of 500 pM virus RNA. This simple, rapid, and direct RNA detection method is expected to provide a complementary method for the detection of viruses mutated at the PCR primer-binding regions for a more precise detection.

KeywordDna Probe Duplex-specific Nuclease Rna Detection Sars-cov-2 Signal Amplification
DOI10.3390/mi12020197
URLView the original
Indexed BySCIE
Language英語English
WOS Research AreaChemistry ; Science & Technology - Other Topics ; Instruments & Instrumentation ; Physics
WOS SubjectChemistry, Analytical ; Nanoscience & Nanotechnology ; Instruments & Instrumentation ; Physics, Applied
WOS IDWOS:000622811600001
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Document TypeJournal article
CollectionINSTITUTE OF MICROELECTRONICS
Faculty of Science and Technology
Corresponding AuthorJia,Yanwei; Martins,Rui P.
Affiliation1.State-Key Laboratory of Analog and Mixed-Signal VLSI,Institute of Microelectronics,University of Macau,Macau,999078,China
2.Faculty of Science and Technology-ECE,University of Macau,Macau,999078,China
3.Faculty of Health Sciences,University of Macau,Macau,999078,China
4.On Leave from Instituto Superior Técnico,Universidade de Lisboa,Lisboa,1049-001,Portugal
First Author AffilicationUniversity of Macau
Corresponding Author AffilicationUniversity of Macau;  Faculty of Science and Technology;  Faculty of Health Sciences
Recommended Citation
GB/T 7714
Liu,Meiqing,Li,Haoran,Jia,Yanwei,et al. SARS-CoV-2 RNA detection with duplex-specific nuclease signal amplification[J]. Micromachines,2021,12(2).
APA Liu,Meiqing,Li,Haoran,Jia,Yanwei,Mak,Pui In,&Martins,Rui P..(2021).SARS-CoV-2 RNA detection with duplex-specific nuclease signal amplification.Micromachines,12(2).
MLA Liu,Meiqing,et al."SARS-CoV-2 RNA detection with duplex-specific nuclease signal amplification".Micromachines 12.2(2021).
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