1. Multifunctional self-priming hairpin probe-based isothermal nucleic acid amplification and its applications for COVID-19 diagnosis
- Author
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Kim, Hansol, Lee, Seoyoung, Ju, Yong, Kim, Hyoyong, Jang, Hyowon, Park, Yeonkyung, Lee, Sang Mo, Yong, Dongeun, Kang, Taejoon, and Park, Hyun Gyu
- Subjects
Analytical Chemistry ,Chemical Sciences ,Biotechnology ,Infectious Diseases ,Coronaviruses ,Emerging Infectious Diseases ,4.2 Evaluation of markers and technologies ,Humans ,Nucleic Acids ,COVID-19 ,COVID-19 Testing ,Nucleic Acid Amplification Techniques ,SARS-CoV-2 ,Biosensing Techniques ,Sensitivity and Specificity ,Isothermal amplification ,Molecular diagnostics ,Self-priming hairpin probe ,Biomedical Engineering ,Nanotechnology ,Bioinformatics ,Analytical chemistry ,Biomedical engineering - Abstract
We herein present a multifunctional self-priming hairpin probe-based isothermal amplification, termed MSH, enabling one-pot detection of target nucleic acids. The sophisticatedly designed multifunctional self-priming hairpin (MSH) probe recognizes the target and rearranges to prime itself, triggering the amplification reaction powered by the continuously repeated extension, nicking, and target recycling. As a consequence, a large number of double-stranded DNA (dsDNA) amplicons are produced that could be monitored in real-time using a dsDNA-intercalating dye. Based on this unique design approach, the nucleocapsid (N) and the open reading frame 1 ab (ORF1ab) genes of SARS-CoV-2 were successfully detected down to 1.664 fM and 0.770 fM, respectively. The practical applicability of our method was validated by accurately diagnosing 60 clinical samples with 93.33% sensitivity and 96.67% specificity. This isothermal one-pot MSH technique holds great promise as a point-of-care testing protocol for the reliable detection of a wide spectrum of pathogens, particularly in resource-limited settings.
- Published
- 2024