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

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...

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Autores principales: Kalita, Manash Jyoti, Dutta, Kalpajit, Hazarika, Gautam, Dutta, Ridip, Kalita, Simanta, Das, Partha Pratim, Sarma, Manash P., Banu, Sofia, Idris, Md. Ghaznavi, Talukdar, Anjan Jyoti, Dutta, Sangitanjan, Sharma, Ajanta, Medhi, Subhash
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8429455/
https://www.ncbi.nlm.nih.gov/pubmed/34504255
http://dx.doi.org/10.1038/s41598-021-97502-1
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author Kalita, Manash Jyoti
Dutta, Kalpajit
Hazarika, Gautam
Dutta, Ridip
Kalita, Simanta
Das, Partha Pratim
Sarma, Manash P.
Banu, Sofia
Idris, Md. Ghaznavi
Talukdar, Anjan Jyoti
Dutta, Sangitanjan
Sharma, Ajanta
Medhi, Subhash
author_facet Kalita, Manash Jyoti
Dutta, Kalpajit
Hazarika, Gautam
Dutta, Ridip
Kalita, Simanta
Das, Partha Pratim
Sarma, Manash P.
Banu, Sofia
Idris, Md. Ghaznavi
Talukdar, Anjan Jyoti
Dutta, Sangitanjan
Sharma, Ajanta
Medhi, Subhash
author_sort Kalita, Manash Jyoti
collection PubMed
description 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.
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spelling pubmed-84294552021-09-10 In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity Kalita, Manash Jyoti Dutta, Kalpajit Hazarika, Gautam Dutta, Ridip Kalita, Simanta Das, Partha Pratim Sarma, Manash P. Banu, Sofia Idris, Md. Ghaznavi Talukdar, Anjan Jyoti Dutta, Sangitanjan Sharma, Ajanta Medhi, Subhash Sci Rep Article 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. Nature Publishing Group UK 2021-09-09 /pmc/articles/PMC8429455/ /pubmed/34504255 http://dx.doi.org/10.1038/s41598-021-97502-1 Text en © The Author(s) 2021 https://creativecommons.org/licenses/by/4.0/Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Article
Kalita, Manash Jyoti
Dutta, Kalpajit
Hazarika, Gautam
Dutta, Ridip
Kalita, Simanta
Das, Partha Pratim
Sarma, Manash P.
Banu, Sofia
Idris, Md. Ghaznavi
Talukdar, Anjan Jyoti
Dutta, Sangitanjan
Sharma, Ajanta
Medhi, Subhash
In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity
title In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity
title_full In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity
title_fullStr In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity
title_full_unstemmed In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity
title_short In-house reverse transcriptase polymerase chain reaction for detection of SARS-CoV-2 with increased sensitivity
title_sort in-house reverse transcriptase polymerase chain reaction for detection of sars-cov-2 with increased sensitivity
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8429455/
https://www.ncbi.nlm.nih.gov/pubmed/34504255
http://dx.doi.org/10.1038/s41598-021-97502-1
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