tetano
Editor, Senior Moderator
Biosens Bioelectron
. 2020 Oct 26;172:112766.
doi: 10.1016/j.bios.2020.112766. Online ahead of print.
opvCRISPR: One-pot visual RT-LAMP-CRISPR platform for SARS-cov-2 detection
Rui Wang[SUP] 1 [/SUP], Chunyan Qian[SUP] 2 [/SUP], Yanan Pang[SUP] 3 [/SUP], Miaomiao Li[SUP] 4 [/SUP], Yu Yang[SUP] 4 [/SUP], Haijing Ma[SUP] 4 [/SUP], Manying Zhao[SUP] 4 [/SUP], Feng Qian[SUP] 4 [/SUP], Hang Yu[SUP] 5 [/SUP], Zhenping Liu[SUP] 6 [/SUP], Ting Ni[SUP] 7 [/SUP], Yan Zheng[SUP] 8 [/SUP], Yongming Wang[SUP] 9 [/SUP]
Affiliations
Abstract
The 2019 novel coronavirus disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected all aspects of human life. Rapid, accurate, sensitive and user friendly detection method is urgently needed to facilitate early intervention and control the spread of SARS-CoV-2. Here, we propose a one-pot visual SARS-CoV-2 detection system named "opvCRISPR" by integrating reverse transcription loop-mediated isothermal amplification (RT-LAMP) and Cas12a cleavage in a single reaction system. We demonstrate that the collateral activity against single-stranded DNA (ssDNA) reporters of activated Cas12a triggered by RT-LAMP amplicon increases detection sensitivity and makes detection results observable with naked eye. The opvCRISPR enables detection at nearly single molecule level in 45 min. We validate this method with 50 SARS-CoV-2 potentially infected clinical samples. The opvCRISPR diagnostic results provide 100% agreement with the Centers for Disease Control and Prevention (CDC)-approved quantitative RT-PCR assay. The opvCRISPR holds great potential for SARS-CoV-2 detection in next-generation point-of-care molecular diagnostics.
Keywords: CRISPR; Molecular diagnosis; One-pot; RT-LAMP; SARS-Cov-2; Visual detection.
. 2020 Oct 26;172:112766.
doi: 10.1016/j.bios.2020.112766. Online ahead of print.
opvCRISPR: One-pot visual RT-LAMP-CRISPR platform for SARS-cov-2 detection
Rui Wang[SUP] 1 [/SUP], Chunyan Qian[SUP] 2 [/SUP], Yanan Pang[SUP] 3 [/SUP], Miaomiao Li[SUP] 4 [/SUP], Yu Yang[SUP] 4 [/SUP], Haijing Ma[SUP] 4 [/SUP], Manying Zhao[SUP] 4 [/SUP], Feng Qian[SUP] 4 [/SUP], Hang Yu[SUP] 5 [/SUP], Zhenping Liu[SUP] 6 [/SUP], Ting Ni[SUP] 7 [/SUP], Yan Zheng[SUP] 8 [/SUP], Yongming Wang[SUP] 9 [/SUP]
Affiliations
- PMID: 33126177
- PMCID: PMC7586109
- DOI: 10.1016/j.bios.2020.112766
Abstract
The 2019 novel coronavirus disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected all aspects of human life. Rapid, accurate, sensitive and user friendly detection method is urgently needed to facilitate early intervention and control the spread of SARS-CoV-2. Here, we propose a one-pot visual SARS-CoV-2 detection system named "opvCRISPR" by integrating reverse transcription loop-mediated isothermal amplification (RT-LAMP) and Cas12a cleavage in a single reaction system. We demonstrate that the collateral activity against single-stranded DNA (ssDNA) reporters of activated Cas12a triggered by RT-LAMP amplicon increases detection sensitivity and makes detection results observable with naked eye. The opvCRISPR enables detection at nearly single molecule level in 45 min. We validate this method with 50 SARS-CoV-2 potentially infected clinical samples. The opvCRISPR diagnostic results provide 100% agreement with the Centers for Disease Control and Prevention (CDC)-approved quantitative RT-PCR assay. The opvCRISPR holds great potential for SARS-CoV-2 detection in next-generation point-of-care molecular diagnostics.
Keywords: CRISPR; Molecular diagnosis; One-pot; RT-LAMP; SARS-Cov-2; Visual detection.