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In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity

Authors :
Simanta Kalita
Manash Pratim Sarma
Sofia Banu
Kalpajit Dutta
Partha Pratim Das
Sangitanjan Dutta
Anjan Jyoti Talukdar
Gautam Hazarika
Ghaznavi Idris
Subhash Medhi
Manash Jyoti Kalita
Ridip Dutta
Ajanta Sharma
Source :
Scientific Reports, Scientific Reports, Vol 11, Iss 1, Pp 1-12 (2021)
Publication Year :
2021
Publisher :
Nature Publishing Group UK, 2021.

Abstract

As the COVID-19 infection continues to ravage the world, the advent of an efficient as well as the economization of the existing RT-PCR based detection assay essentially can become a blessing in these testing times and significantly help in the management of the pandemic. This study demonstrated an innovative and rapid corroboration of COVID-19 test based on innovative multiplex PCR. An assessment of optimal PCR conditions to simultaneously amplify the SARS-CoV-2 genes E, S and RdRp has been made by fast-conventional and HRM coupled multiplex real-time PCR using the same sets of primers. All variables of practical value were studied by amplifying known target-sequences from ten-fold dilutions of archived positive samples of COVID-19 disease. The multiplexing with newly designed E, S and RdRp primers have shown an efficient amplification of the target region of SARS-CoV-2. A distinct amplification was observed in 37 min using thermal cycler while it took 96 min in HRM coupled real time detection using SYBR green over a wide range of template concentrations. Our findings revealed decent concordance with other commercially available detection kits. This fast HRM coupled multiplex real-time PCR with SYBR green approach offers rapid and sensitive detection of SARS-CoV-2 in a cost-effective manner apart from the added advantage of primer compatibility for use in conventional multiplex PCR. The highly reproducible novel approach can propel extended applicability for developing sustainable commercial product besides providing relief to a resource limited setting.

Details

Language :
English
ISSN :
20452322
Volume :
11
Database :
OpenAIRE
Journal :
Scientific Reports
Accession number :
edsair.doi.dedup.....23cf724abb01bc4a8cd8dc496af68cbc