Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the <i>N</i> Gene of SARS-CoV-2
COVID-19 was officially declared a global pandemic disease on 11 March 2020, with severe implications for healthcare systems, economic activity, and human life worldwide. Fast and sensitive amplification of the severe acute respiratory syndrome virus 2 (SARS-CoV-2) nucleic acids is critical for cont...
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| Format: | Article |
| Language: | English |
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MDPI AG
2022-12-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/23/23/15269 |
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| author | Huan Cui Fei Tu Cheng Zhang Chunmao Zhang Kui Zhao Juxiang Liu Shishan Dong Ligong Chen Jun Liu Zhendong Guo |
| author_facet | Huan Cui Fei Tu Cheng Zhang Chunmao Zhang Kui Zhao Juxiang Liu Shishan Dong Ligong Chen Jun Liu Zhendong Guo |
| author_sort | Huan Cui |
| collection | DOAJ |
| description | COVID-19 was officially declared a global pandemic disease on 11 March 2020, with severe implications for healthcare systems, economic activity, and human life worldwide. Fast and sensitive amplification of the severe acute respiratory syndrome virus 2 (SARS-CoV-2) nucleic acids is critical for controlling the spread of this disease. Here, a real-time reverse transcription recombinase-aided amplification (RT-RAA) assay, targeting conserved positions in the nucleocapsid protein gene (<i>N</i> gene) of SARS-CoV-2, was successfully established for SARS-CoV-2. The assay was specific to SARS-CoV-2, and there was no cross-reaction with other important viruses. The sensitivity of the real-time RT-RAA assay was 142 copies per reaction at 95% probability. Furthermore, 100% concordance between the real-time RT-RAA and RT-qPCR assays was achieved after testing 72 clinical specimens. Further linear regression analysis indicated a significant correlation between the real-time RT-RAA and RT-qPCR assays with an R2 value of 0.8149 (<i>p</i> < 0.0001). In addition, the amplicons of the real-time RT-RAA assay could be directly visualized by a portable blue light instrument, making it suitable for the rapid amplification of SARS-CoV-2 in resource-limited settings. Therefore, the real-time RT-RAA method allows the specific, sensitive, simple, rapid, and reliable detection of SARS-CoV-2. |
| first_indexed | 2024-03-09T17:44:28Z |
| format | Article |
| id | doaj.art-5b88300e055642a9bcad66f8e7671f5f |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-09T17:44:28Z |
| publishDate | 2022-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-5b88300e055642a9bcad66f8e7671f5f2023-11-24T11:16:08ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-12-0123231526910.3390/ijms232315269Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the <i>N</i> Gene of SARS-CoV-2Huan Cui0Fei Tu1Cheng Zhang2Chunmao Zhang3Kui Zhao4Juxiang Liu5Shishan Dong6Ligong Chen7Jun Liu8Zhendong Guo9Changchun Veterinary Research Institute, Chinese Academy of Agriculture Sciences, Changchun 130122, ChinaChangchun Veterinary Research Institute, Chinese Academy of Agriculture Sciences, Changchun 130122, ChinaCollege of Veterinary Medicine, Hebei Agricultural University, Baoding 071000, ChinaChangchun Veterinary Research Institute, Chinese Academy of Agriculture Sciences, Changchun 130122, ChinaCollege of Animal Medicine, Jilin University, Changchun 130062, ChinaCollege of Veterinary Medicine, Hebei Agricultural University, Baoding 071000, ChinaCollege of Veterinary Medicine, Hebei Agricultural University, Baoding 071000, ChinaCollege of Veterinary Medicine, Hebei Agricultural University, Baoding 071000, ChinaChangchun Veterinary Research Institute, Chinese Academy of Agriculture Sciences, Changchun 130122, ChinaChangchun Veterinary Research Institute, Chinese Academy of Agriculture Sciences, Changchun 130122, ChinaCOVID-19 was officially declared a global pandemic disease on 11 March 2020, with severe implications for healthcare systems, economic activity, and human life worldwide. Fast and sensitive amplification of the severe acute respiratory syndrome virus 2 (SARS-CoV-2) nucleic acids is critical for controlling the spread of this disease. Here, a real-time reverse transcription recombinase-aided amplification (RT-RAA) assay, targeting conserved positions in the nucleocapsid protein gene (<i>N</i> gene) of SARS-CoV-2, was successfully established for SARS-CoV-2. The assay was specific to SARS-CoV-2, and there was no cross-reaction with other important viruses. The sensitivity of the real-time RT-RAA assay was 142 copies per reaction at 95% probability. Furthermore, 100% concordance between the real-time RT-RAA and RT-qPCR assays was achieved after testing 72 clinical specimens. Further linear regression analysis indicated a significant correlation between the real-time RT-RAA and RT-qPCR assays with an R2 value of 0.8149 (<i>p</i> < 0.0001). In addition, the amplicons of the real-time RT-RAA assay could be directly visualized by a portable blue light instrument, making it suitable for the rapid amplification of SARS-CoV-2 in resource-limited settings. Therefore, the real-time RT-RAA method allows the specific, sensitive, simple, rapid, and reliable detection of SARS-CoV-2.https://www.mdpi.com/1422-0067/23/23/15269reverse transcription recombinase-aided amplification (RT-RAA)SARS-CoV-2isothermal amplificationrapid diagnosisvisual amplification |
| spellingShingle | Huan Cui Fei Tu Cheng Zhang Chunmao Zhang Kui Zhao Juxiang Liu Shishan Dong Ligong Chen Jun Liu Zhendong Guo Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the <i>N</i> Gene of SARS-CoV-2 International Journal of Molecular Sciences reverse transcription recombinase-aided amplification (RT-RAA) SARS-CoV-2 isothermal amplification rapid diagnosis visual amplification |
| title | Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the <i>N</i> Gene of SARS-CoV-2 |
| title_full | Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the <i>N</i> Gene of SARS-CoV-2 |
| title_fullStr | Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the <i>N</i> Gene of SARS-CoV-2 |
| title_full_unstemmed | Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the <i>N</i> Gene of SARS-CoV-2 |
| title_short | Real-Time Reverse Transcription Recombinase-Aided Amplification Assay for Rapid Amplification of the <i>N</i> Gene of SARS-CoV-2 |
| title_sort | real time reverse transcription recombinase aided amplification assay for rapid amplification of the i n i gene of sars cov 2 |
| topic | reverse transcription recombinase-aided amplification (RT-RAA) SARS-CoV-2 isothermal amplification rapid diagnosis visual amplification |
| url | https://www.mdpi.com/1422-0067/23/23/15269 |
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