A Simple and Multiplex Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of SARS-CoV
Abstract The current diagnosis of severe acute respiratory syndrome-associated coronavirus (SARS-CoV) relies on laboratory-based tests since its clinical features are nonspecific, unlike other respiratory pathogens. Therefore, the development of a rapid and simple method for on-site detection of SARS-CoV is crucial for the identification and prevention of future SARS outbreaks. In this study, a simple colorimetric and multiplex loop-mediated isothermal amplification (LAMP) assay was developed to rapid screening of severe acute respiratory syndrome-associated coronavirus (SARS-CoV). It can be visually detected based on color change and monitored in real-time with fluorescent signals. The performance of this assay, based on six primers targeting open reading frame (ORF1b) and nucleocapsid (N) genes located in different regions of the SARS-CoV, was compared with real-time RT-PCR assay using various concentrations of target genes. The detection limit of the LAMP assay was comparable to that of real-time RT-PCR assay and therefore a few target RNA to $ 10^{4} $-$ 10^{5} $ copies could be detected within a short period of time (20–25 min). In addition, we established a multiplex real-time LAMP assay to simultaneously detect two target regions within the SARS-CoV genome. Two target sequences were amplified by specific primers in the same reaction tube and revealed that it was able to detect down to $ 10^{5} $ copies. The standard curve had a linear relationship with similar amplification efficiencies. The LAMP assay results in shorter “sample-to-answer” time than conventional PCR method. Therefore, it is suitable not only for diagnosis of clinical test, but also for surveillance of SARS virus in developing countries..
| Medienart: |
Artikel |
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| Erscheinungsjahr: |
2019 |
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| Erschienen: |
2019 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:13 |
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| Enthalten in: |
BioChip journal - 13(2019), 4 vom: 11. Nov., Seite 341-351 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Kim, Jin Hwa [VerfasserIn] |
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| Links: |
Volltext [lizenzpflichtig] |
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| Themen: |
Colorimetric detection |
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| Anmerkungen: |
© The Korean BioChip Society and Springer 2019 |
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| doi: |
10.1007/s13206-019-3404-3 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
OLC2028446692 |
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| 520 | |a Abstract The current diagnosis of severe acute respiratory syndrome-associated coronavirus (SARS-CoV) relies on laboratory-based tests since its clinical features are nonspecific, unlike other respiratory pathogens. Therefore, the development of a rapid and simple method for on-site detection of SARS-CoV is crucial for the identification and prevention of future SARS outbreaks. In this study, a simple colorimetric and multiplex loop-mediated isothermal amplification (LAMP) assay was developed to rapid screening of severe acute respiratory syndrome-associated coronavirus (SARS-CoV). It can be visually detected based on color change and monitored in real-time with fluorescent signals. The performance of this assay, based on six primers targeting open reading frame (ORF1b) and nucleocapsid (N) genes located in different regions of the SARS-CoV, was compared with real-time RT-PCR assay using various concentrations of target genes. The detection limit of the LAMP assay was comparable to that of real-time RT-PCR assay and therefore a few target RNA to $ 10^{4} $-$ 10^{5} $ copies could be detected within a short period of time (20–25 min). In addition, we established a multiplex real-time LAMP assay to simultaneously detect two target regions within the SARS-CoV genome. Two target sequences were amplified by specific primers in the same reaction tube and revealed that it was able to detect down to $ 10^{5} $ copies. The standard curve had a linear relationship with similar amplification efficiencies. The LAMP assay results in shorter “sample-to-answer” time than conventional PCR method. Therefore, it is suitable not only for diagnosis of clinical test, but also for surveillance of SARS virus in developing countries. | ||
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| 700 | 1 | |a Chung, Doo Ryeon |4 aut | |
| 700 | 1 | |a Kim, Jiyeon |4 aut | |
| 700 | 1 | |a Hwang, Eung Soo |4 aut | |
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