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

Autor: Kalita MJ; Laboratory of Molecular Virology and Oncology, Department of Bioengineering and Technology, Gauhati University, Guwahati, Assam, 781014, India., Dutta K; Laboratory of Molecular Virology and Oncology, Department of Bioengineering and Technology, Gauhati University, Guwahati, Assam, 781014, India., Hazarika G; Laboratory of Molecular Virology and Oncology, Department of Bioengineering and Technology, Gauhati University, Guwahati, Assam, 781014, India., Dutta R; Department of Microbiology, GMCH, Guwahati, Assam, 781032, India., Kalita S; Laboratory of Molecular Virology and Oncology, Department of Bioengineering and Technology, Gauhati University, Guwahati, Assam, 781014, India.; Department of Medicine, GMCH, Guwahati, Assam, 781032, India., Das PP; Laboratory of Molecular Virology and Oncology, Department of Bioengineering and Technology, Gauhati University, Guwahati, Assam, 781014, India.; Department of Medicine, GMCH, Guwahati, Assam, 781032, India., Sarma MP; Department of Biotechnology, Assam Down Town University, Guwahati, Assam, 781068, India., Banu S; Laboratory of Molecular Virology and Oncology, Department of Bioengineering and Technology, Gauhati University, Guwahati, Assam, 781014, India., Idris MG; Laboratory of Molecular Virology and Oncology, Department of Bioengineering and Technology, Gauhati University, Guwahati, Assam, 781014, India., Talukdar AJ; Department of Medicine, GMCH, Guwahati, Assam, 781032, India., Dutta S; Department of Medicine, GMCH, Guwahati, Assam, 781032, India., Sharma A; Department of Microbiology, GMCH, Guwahati, Assam, 781032, India. ajantasharma2002@yahoo.com., Medhi S; Laboratory of Molecular Virology and Oncology, Department of Bioengineering and Technology, Gauhati University, Guwahati, Assam, 781014, India. subhashmedhi@gauhati.ac.in.
Jazyk: angličtina
Zdroj: Scientific reports [Sci Rep] 2021 Sep 09; Vol. 11 (1), pp. 17878. Date of Electronic Publication: 2021 Sep 09.
DOI: 10.1038/s41598-021-97502-1
Abstrakt: 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.
(© 2021. The Author(s).)
Databáze: MEDLINE
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