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.
Artículo
en Inglés
| 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.
Palabras clave
Texto completo:
Disponible
Colección:
Bases de datos internacionales
Base de datos:
MEDLINE
Asunto principal:
ADN Viral
/
ARN Viral
/
Estadística como Asunto
/
Técnicas de Amplificación de Ácido Nucleico
/
Recombinasas
/
ADN Polimerasa Dirigida por ADN
/
SARS-CoV-2
Tipo de estudio:
Estudios diagnósticos
/
Estudio experimental
Idioma:
Inglés
Revista:
Biochem Biophys Res Commun
Año:
2021
Tipo del documento:
Artículo
País de afiliación:
J.bbrc.2021.06.023
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