Optimization of reaction condition of recombinase polymerase amplification to detect SARS-CoV-2 DNA and RNA using a statistical method.
Biochem Biophys Res Commun
; 567: 195-200, 2021 08 27.
Article
in English
| MEDLINE | ID: covidwho-1263226
ABSTRACT
Recombinase polymerase amplification (RPA) is an isothermal reaction that amplifies a target DNA sequence with a recombinase, a single-stranded DNA-binding protein (SSB), and a strand-displacing DNA polymerase. In this study, we optimized the reaction conditions of RPA to detect SARS-CoV-2 DNA and RNA using a statistical method to enhance the sensitivity. In vitro synthesized SARS-CoV-2 DNA and RNA were used as targets. After evaluating the concentration of each component, the uvsY, gp32, and ATP concentrations appeared to be rate-determining factors. In particular, the balance between the binding and dissociation of uvsX and DNA primer was precisely adjusted. Under the optimized condition, 60 copies of the target DNA were specifically detected. Detection of 60 copies of RNA was also achieved. Our results prove the fabrication flexibility of RPA reagents, leading to an expansion of the use of RPA in various fields.
Keywords
Full text:
Available
Collection:
International databases
Database:
MEDLINE
Main subject:
DNA, Viral
/
RNA, Viral
/
Statistics as Topic
/
Nucleic Acid Amplification Techniques
/
Recombinases
/
DNA-Directed DNA Polymerase
/
SARS-CoV-2
Type of study:
Diagnostic study
/
Experimental Studies
Language:
English
Journal:
Biochem Biophys Res Commun
Year:
2021
Document Type:
Article
Affiliation country:
J.bbrc.2021.06.023
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