A high-specificity flap probe-based isothermal nucleic acid amplification method based on recombinant FEN1-Bst DNA polymerase.
Biosens Bioelectron
; 192: 113503, 2021 Nov 15.
Article
in English
| MEDLINE | ID: covidwho-1309167
ABSTRACT
The COVID-19 pandemic has unfortunately demonstrated how easily infectious diseases can spread and harm human life and society. As of writing, pandemic has now been on-going for more than one year. There is an urgent need for new nucleic acid-based methods that can be used to diagnose pathogens early, quickly, and accurately to effectively impede the spread of infections and gain control of epidemics. We developed a flap probe-based isothermal nucleic acid amplification method that is triggered by recombinant FEN1-Bst DNA polymerase, which-through enzymatic engineering-has both DNA synthesis, strand displacement and cleavage functions. This novel method offers a simpler and more specific probe-primer pair than those of other isothermal amplifications. We tested the method's ability to detect SARS-CoV-2 (both ORF1ab and N genes), rotavirus, and Chlamydia trachomatis. The limits of detection were 10 copies/µL for rotavirus, C. trachomatis, and SARS-CoV-2 N gene, and 100 copies/µL for SARS-CoV-2 ORF1ab gene. There were no cross-reactions among 11 other common pathogens with characteristics similar to those of the test target, and the method showed 100% sensitivity and 100% specificity in clinical comparisons with RT-PCR testing. In addition to real-time detection, the endpoint could be displayed under a transilluminator, which is a convenient reporting method for point-of-care test settings. Therefore, this novel nucleic acid senor has great potential for use in clinical diagnostics, epidemic prevention, and epidemic control.
Keywords
Full text:
Available
Collection:
International databases
Database:
MEDLINE
Main subject:
Biosensing Techniques
/
Chlamydia trachomatis
/
Rotavirus
/
SARS-CoV-2
Type of study:
Diagnostic study
/
Prognostic study
/
Randomized controlled trials
Limits:
Humans
Language:
English
Journal:
Biosens Bioelectron
Journal subject:
Biotechnology
Year:
2021
Document Type:
Article
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