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URL: https://dx.doi.org/DOI:10.17504/protocols.io.bhjgj4jw

Authors: Sureshnee Pillay, Jennifer Giandhari, Houriiyah Tegally, Eduan Wilkinson, Benjamin Chimukangara, Richard Lessells, Yunus Moosa, Inbal Gazy, Maryam Fish, Lavanya Singh, Khulekani Sedwell Khanyile, Vagner Fonseca, Marta Giovanetti, Luiz Carols Alcantara, Tulio de Oliveira, Jennifer Giandhari

Group: Coronavirus Method Development Community, KRISP

Summary: This protocol describes the procedure for generating cDNA from SARS-CoV-2 viral nucleic acid extracts and subsequently producing amplicons tiling the viral genome sequencing. It uses the V3 nCov-2019 primers from the ARTIC network. This is followed by library construction using Nextera Flex, which we found to save 9h of hands on time as compared with original protocol that use TrueSeq for library contstruction. It also describes the pooling of samples and quantitation, prior to sequencing on the Illumina Miseq and NextSeq. It is adapted from the nCov-2019 sequencing protocol from Quick and colleagues, which can be found here:

Associated Publications: Pillay S, Giandhari J, Tegally H, Wilkinson E, Chimukangara B, Lessells R, Moosa Y, Mattison S, Gazy I, Fish M, Singh L, Khanyile KS, San JE, Fonseca V, Giovanetti M, Alcantara LC, Oliveira Td, Whole Genome Sequencing of SARS-CoV-2: Adapting Illumina Protocols for Quick and Accurate Outbreak Investigation during a Pandemic. Genes 11(8). doi: 10.3390/genes11080949

Affiliations: KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa; Infectious Diseases Department, Nelson R Mandela School of Medicine, University of KwaZulu-Natal, Durban, South Africa, Infectious Diseases Department, Nelson R Mandela School of Medicine, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa; Laboratorio de Genetica Celular e Molecular, ICB, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil; Laboratório de Flavivírus, Instituto Oswaldo Cruz Fiocruz, Rio de Janeiro, Brazil, Laboratório de Flavivírus, Instituto Oswaldo Cruz Fiocruz, Rio de Janeiro, Brazil, Laboratorio de Genetica Celular e Molecular, ICB, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil; Laboratório de Flavivírus, Instituto Oswaldo Cruz Fiocruz, Rio de Janeiro, Brazil, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa; Centre for Aids Programme of Research in South Africa (CAPRISA), Durban, South Africa; Department of Global Health, University of Washington, Seattle, Washington, USA, KwaZulu-Natal Research Innovation and Sequencing Platform (KRISP), School of Laboratory Medicine & Medical Sciences, University of KwaZulu-Natal, Durban, South Africa; Centre for Aids Programme of Research in South Africa (CAPRISA), Durban, South Africa; Department of Global Health, University of Washington, Seattle, Washington, USA

External URL: https://www.krisp.org.za/publications.php?pubid=292

Version: 1

Publication Date: 2020

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