A Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2
Abstract
:1. Introduction
2. Results
2.1. Primer Design
2.2. Specificity and Sensitivity of the SARS-CoV-2 RT-LAMP Assay
2.3. Visual Detection
2.4. Evaluation of the Novel SARS-CoV-2 RT-LAMP Assay Using Clinical Samples
3. Discussion
4. Materials and Methods
4.1. RNA Extraction
4.2. Reaction System of the Novel RT-LAMP Assay
4.3. Specificity of the Novel RT-LAMP Assay
4.4. Sensitivity and Limit of Detection (LOD)
4.5. Evaluation of the Novel SARS-CoV-2 Detection Assay Using Clinical Samples
4.6. Visual Detection
4.7. Ethics Statement
5. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
COVID-19 | Coronavirus disease 2019 |
SARS-CoV | Severe Acute Respiratory Syndrome Coronavirus |
MERS-CoV | Middle East respiratory syndrome coronavirus |
LAMP | Loop-mediated isothermal amplification |
RSV | Respiratory syncytial virus |
ACE2 | Angiotensin converting enzyme II |
POCT | Point-of-Care testing |
BALF | Bronchoalveolar lavage fluid |
NCBI | National Centre for Biotechnology Information |
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Primers | Sequence (5’-3’) | Length (nt) |
---|---|---|
F3 | GCCAAAAGGCTTCTACGCA | 19 |
B3 | TTGCTCTCAAGCTGGTTCAA | 20 |
FIP | TCCCCTACTGCTGCCTGGAGCAGTCAAGCCTCTTCTCGTT | 40 |
BIP | TCTCCTGCTAGAATGGCTGGCATCTGTCAAGCAGCAGCAAAG | 42 |
LB | TGGCGGTGATGCTGCTCTT | 19 |
Dilution | Standard (Copies/Reaction) | Positive/Total Tested |
---|---|---|
1× | 1963 | 10/10 |
5× | 393 | 10/10 |
5× | 79 | 8/10 |
5× | 16 | 6/10 |
5× | 3 | 2/10 |
LOD | 118.6 copies/reaction |
RT-qPCR Assay | The novel RT-LAMP Assay | Total | Positive Rate (%) | Concordance Rate (%) | |
---|---|---|---|---|---|
Positive | Negative | ||||
Positive | 34 | 2 | 36 | 64.3% | 92.9% |
negative | 2 | 18 | 20 | ||
Total | 36 | 20 | 56 | ||
Positive rate (%) | 63.4% |
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Lu, R.; Wu, X.; Wan, Z.; Li, Y.; Jin, X.; Zhang, C. A Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2. Int. J. Mol. Sci. 2020, 21, 2826. https://doi.org/10.3390/ijms21082826
Lu R, Wu X, Wan Z, Li Y, Jin X, Zhang C. A Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2. International Journal of Molecular Sciences. 2020; 21(8):2826. https://doi.org/10.3390/ijms21082826
Chicago/Turabian StyleLu, Renfei, Xiuming Wu, Zhenzhou Wan, Yingxue Li, Xia Jin, and Chiyu Zhang. 2020. "A Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2" International Journal of Molecular Sciences 21, no. 8: 2826. https://doi.org/10.3390/ijms21082826
APA StyleLu, R., Wu, X., Wan, Z., Li, Y., Jin, X., & Zhang, C. (2020). A Novel Reverse Transcription Loop-Mediated Isothermal Amplification Method for Rapid Detection of SARS-CoV-2. International Journal of Molecular Sciences, 21(8), 2826. https://doi.org/10.3390/ijms21082826