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Short report
Lucy Baldeón
Central University of Ecuador
Corresponding Author
ORCiD: https://orcid.org/0000-0002-0447-0136
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The World Health Organization (WHO) reported about 30 million cases of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, and around 1 million deaths worldwide. Ecuador is the country with the highest mortality rate of confirmed cases in South America where both the poor ability to identify SARS-CoV-2 carriers and shortages in reagent supply have contributed the high infection rate observed. Hence, there is an urgent need to develop, standardize and validate an in-house protocol that cannot only reduces testing costs, but also increase the ability to screen widely the population. Primer-probe sets for the SARS-CoV-2 envelope protein E and the human ribonuclease P (RP) were validated for a duplex quantitative Reverse Transcription Polymerase Chain Reaction (RT-qPCR) for the Coronavirus Disease-19 (COVID-19) detection. Optimal E primers concentration was 400 nM and TaqMan probe 200 nM. The primer efficiency was set at 94.9% and R2 value at 0.99, which was comparable to commercial kits. The lower detection limit was found at 15 copies/ μL (50 copies/rx). In comparison to a Loop-mediated Isothermal Amplification (LAMP) commercial kit, there was a higher detection rate (30%) and results were highly reproducible (95%). We were able to develop a highly sensitive and low-cost duplex in-house RT-qPCR test for COVID-19 detection comparable to other commercially available kits.
Keywords
in-house, RT-qPCR, SARS-CoV-2, gene E, COVID-19
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A new preprint design is coming!
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This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Short report
Lucy Baldeón
Central University of Ecuador
Corresponding Author
ORCiD: https://orcid.org/0000-0002-0447-0136
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 15 Oct, 2020
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
The World Health Organization (WHO) reported about 30 million cases of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, and around 1 million deaths worldwide. Ecuador is the country with the highest mortality rate of confirmed cases in South America where both the poor ability to identify SARS-CoV-2 carriers and shortages in reagent supply have contributed the high infection rate observed. Hence, there is an urgent need to develop, standardize and validate an in-house protocol that cannot only reduces testing costs, but also increase the ability to screen widely the population. Primer-probe sets for the SARS-CoV-2 envelope protein E and the human ribonuclease P (RP) were validated for a duplex quantitative Reverse Transcription Polymerase Chain Reaction (RT-qPCR) for the Coronavirus Disease-19 (COVID-19) detection. Optimal E primers concentration was 400 nM and TaqMan probe 200 nM. The primer efficiency was set at 94.9% and R2 value at 0.99, which was comparable to commercial kits. The lower detection limit was found at 15 copies/ μL (50 copies/rx). In comparison to a Loop-mediated Isothermal Amplification (LAMP) commercial kit, there was a higher detection rate (30%) and results were highly reproducible (95%). We were able to develop a highly sensitive and low-cost duplex in-house RT-qPCR test for COVID-19 detection comparable to other commercially available kits.
Figure 1
Figure 2
Figure 3
Figure 4
This is a list of supplementary files associated with this preprint. Click to download.
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