Novel reverse transcription-multiple inner primer loop-mediated isothermal amplification (RT-MIPLAMP) for visual and sensitive detection of SARS-CoV-2

Since the end of 2019, outbreaks of COVID-19 pandemics have continued in different areas worldwide, which exacerbates the need for rapid, sensitive and simple methods for diagnosis. Currently, COVID-19 diagnosis mainly relies on reverse transcription-polymerase chain reaction (RT-PCR), which require...

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Bibliographic Details
Published in:Analytical methods 2022-12, Vol.14 (47), p.512-518
Main Authors: He, Xiaofei, Su, Fengxia, Chen, Yutong, Li, Zhengping
Format: Article
Language:English
Subjects:
Coronaviruses
COVID-19
COVID-19 - diagnosis
COVID-19 Testing
Diagnosis
Gene amplification
Humans
N gene
Pandemics
Polymerase chain reaction
Reaction time
Reverse Transcription
Saliva
SARS-CoV-2 - genetics
Severe acute respiratory syndrome coronavirus 2
Viral diseases
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Summary:Since the end of 2019, outbreaks of COVID-19 pandemics have continued in different areas worldwide, which exacerbates the need for rapid, sensitive and simple methods for diagnosis. Currently, COVID-19 diagnosis mainly relies on reverse transcription-polymerase chain reaction (RT-PCR), which requires sophisticated instruments. Reverse transcription-loop mediated isothermal amplification (RT-LAMP), due to its isothermal nature and high specificity, can be used as an alternative. In this paper, a novel visual reverse transcription-multiple inner primer loop-mediated isothermal amplification (RT-MIPLAMP) method is established based on RT-LAMP by adding a pair of inner primers. The RT-MIPLAMP method has a higher sensitivity and shorter reaction time compared with conventional RT-LAMP. By using RT-MIPLAMP, as low as 6 × 10 3 copies per mL in vitro transcribed (IVT) N gene can be detected within 55 min. Meanwhile, as low as 6 × 10 4 copies per mL IVT N gene is detectable with conventional RT-LAMP within 80 min. The feasibility of visual RT-MIPLAMP is also validated by detecting the N gene spiked into one healthy volunteer's saliva and the full-length RNA in pseudoviruses, indicating the great potential of visual RT-MIPLAMP for SARS-CoV-2 identification. A novel reverse transcription-multiple inner primer loop-mediated isothermal amplification (RT-MIPLAMP) method is developed for sensitive detection of SARS-CoV-2.
ISSN:1759-9660
1759-9679
DOI:10.1039/d2ay01330d