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One-step RNA extraction for RT-qPCR detection of 2019-nCoV

Monica Sentmanat, Evguenia Kouranova, Xiaoxia Cui
doi: https://doi.org/10.1101/2020.04.02.022384
Monica Sentmanat
Genome Engineering & iPSC Center (GEiC), Department of Genetics, Washington University in St. Louis School of Medicine, 660 S. Euclid Ave., St. Louis, MO 63110
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Evguenia Kouranova
Genome Engineering & iPSC Center (GEiC), Department of Genetics, Washington University in St. Louis School of Medicine, 660 S. Euclid Ave., St. Louis, MO 63110
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Xiaoxia Cui
Genome Engineering & iPSC Center (GEiC), Department of Genetics, Washington University in St. Louis School of Medicine, 660 S. Euclid Ave., St. Louis, MO 63110
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  • For correspondence: x.cui@wustl.edu
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ABSTRACT

The global outbreak of coronavirus disease 2019 (COVID-19) has placed an unprecedented burden on healthcare systems as the virus spread from the initial 27 reported cases in the city of Wuhan, China to a global pandemic in under three months1. Resources essential to monitoring virus transmission have been challenged with a demand for expanded surveillance. The CDC 2019-nCoV Real-Time Diagnostic Panel uses a real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR) consisting of two TaqMan probe and primer sets specific for the 2019-nCoV N gene, which codes for the nucleocapsid structural protein that encapsulates viral RNA, for the qualitative detection of 2019-nCoV viral RNA in respiratory samples. To isolate RNA from respiratory samples, the CDC lists RNA extraction kits from three manufacturers. In anticipation of a limited supply chain of RNA extraction kits and the need for test scalability, we sought to identify alternative RNA extraction methods. Here we show that direct lysis of respiratory samples can be used in place of RNA extraction kits to run the CDC 2019-nCoV Real-Time Diagnostic assay with the additional benefits of higher throughput, lower cost, faster turnaround and possibly higher senitivity and improved saftey.

Competing Interest Statement

The authors have declared no competing interest.

Footnotes

  • Positive control description has been corrected. A DNA, not RNA template was used for positive control counterparts described in the Results section.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted April 08, 2020.
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One-step RNA extraction for RT-qPCR detection of 2019-nCoV
Monica Sentmanat, Evguenia Kouranova, Xiaoxia Cui
bioRxiv 2020.04.02.022384; doi: https://doi.org/10.1101/2020.04.02.022384
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One-step RNA extraction for RT-qPCR detection of 2019-nCoV
Monica Sentmanat, Evguenia Kouranova, Xiaoxia Cui
bioRxiv 2020.04.02.022384; doi: https://doi.org/10.1101/2020.04.02.022384

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