A Novel Approach to the Bioluminescent Detection of the SARS-CoV-2 ORF1ab Gene by Coupling Isothermal RNA Reverse Transcription Amplification with a Digital PCR Approach
The COVID-19 pandemic caused by the SARS-CoV-2 virus, which first emerged in December 2019, represents an ongoing global public health emergency. Here, we developed an improved and highly sensitive approach to SARS-CoV-2 detection via coupling bioluminescence in real-time (BART) and reverse-transcri...
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-01-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/22/3/1017 |
| _version_ | 1797409254873759744 |
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| author | Zhongjie Fei Rongbin Wei Chu Cheng Pengfeng Xiao |
| author_facet | Zhongjie Fei Rongbin Wei Chu Cheng Pengfeng Xiao |
| author_sort | Zhongjie Fei |
| collection | DOAJ |
| description | The COVID-19 pandemic caused by the SARS-CoV-2 virus, which first emerged in December 2019, represents an ongoing global public health emergency. Here, we developed an improved and highly sensitive approach to SARS-CoV-2 detection via coupling bioluminescence in real-time (BART) and reverse-transcriptase loop-mediated amplification (RT-LAMP) protocols (RT-LAMP-BART) and was also compatible with a digital LAMP system (Rainsuit), which did not allow for real-time quantification but did, nonetheless, facilitate absolute quantification with a comparable detection limit of 10<sup>4</sup> copies/mL. Through improving RNA availability in samples to ensure the target RNA present in reaction, we additionally developed a simulated digital RT-LAMP approach using this same principle to enlarge the overall reaction volume and to achieve real-time detection with a limit of detection of 10 copies/mL, and with further improvements in the overall dynamic range of this assay system being achieved through additional optimization. |
| first_indexed | 2024-03-09T04:11:41Z |
| format | Article |
| id | doaj.art-681c974cefb34b0bbcf7578105e74c8c |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-09T04:11:41Z |
| publishDate | 2021-01-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-681c974cefb34b0bbcf7578105e74c8c2023-12-03T13:59:16ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-01-01223101710.3390/ijms22031017A Novel Approach to the Bioluminescent Detection of the SARS-CoV-2 ORF1ab Gene by Coupling Isothermal RNA Reverse Transcription Amplification with a Digital PCR ApproachZhongjie Fei0Rongbin Wei1Chu Cheng2Pengfeng Xiao3State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, ChinaState Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, ChinaState Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, ChinaState Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, ChinaThe COVID-19 pandemic caused by the SARS-CoV-2 virus, which first emerged in December 2019, represents an ongoing global public health emergency. Here, we developed an improved and highly sensitive approach to SARS-CoV-2 detection via coupling bioluminescence in real-time (BART) and reverse-transcriptase loop-mediated amplification (RT-LAMP) protocols (RT-LAMP-BART) and was also compatible with a digital LAMP system (Rainsuit), which did not allow for real-time quantification but did, nonetheless, facilitate absolute quantification with a comparable detection limit of 10<sup>4</sup> copies/mL. Through improving RNA availability in samples to ensure the target RNA present in reaction, we additionally developed a simulated digital RT-LAMP approach using this same principle to enlarge the overall reaction volume and to achieve real-time detection with a limit of detection of 10 copies/mL, and with further improvements in the overall dynamic range of this assay system being achieved through additional optimization.https://www.mdpi.com/1422-0067/22/3/1017SARS-CoV-2accurate and rapid identificationRT-LAMP-BARTdigital PCR systemdigital RT-LAMP approach |
| spellingShingle | Zhongjie Fei Rongbin Wei Chu Cheng Pengfeng Xiao A Novel Approach to the Bioluminescent Detection of the SARS-CoV-2 ORF1ab Gene by Coupling Isothermal RNA Reverse Transcription Amplification with a Digital PCR Approach International Journal of Molecular Sciences SARS-CoV-2 accurate and rapid identification RT-LAMP-BART digital PCR system digital RT-LAMP approach |
| title | A Novel Approach to the Bioluminescent Detection of the SARS-CoV-2 ORF1ab Gene by Coupling Isothermal RNA Reverse Transcription Amplification with a Digital PCR Approach |
| title_full | A Novel Approach to the Bioluminescent Detection of the SARS-CoV-2 ORF1ab Gene by Coupling Isothermal RNA Reverse Transcription Amplification with a Digital PCR Approach |
| title_fullStr | A Novel Approach to the Bioluminescent Detection of the SARS-CoV-2 ORF1ab Gene by Coupling Isothermal RNA Reverse Transcription Amplification with a Digital PCR Approach |
| title_full_unstemmed | A Novel Approach to the Bioluminescent Detection of the SARS-CoV-2 ORF1ab Gene by Coupling Isothermal RNA Reverse Transcription Amplification with a Digital PCR Approach |
| title_short | A Novel Approach to the Bioluminescent Detection of the SARS-CoV-2 ORF1ab Gene by Coupling Isothermal RNA Reverse Transcription Amplification with a Digital PCR Approach |
| title_sort | novel approach to the bioluminescent detection of the sars cov 2 orf1ab gene by coupling isothermal rna reverse transcription amplification with a digital pcr approach |
| topic | SARS-CoV-2 accurate and rapid identification RT-LAMP-BART digital PCR system digital RT-LAMP approach |
| url | https://www.mdpi.com/1422-0067/22/3/1017 |
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