Automated SARS-COV-2 RNA extraction from patient nasopharyngeal samples using a modified DNA extraction kit for high throughput testing.

Autor:	Al-Saud H; From the Saudi Human Genome Project, King Abdulaziz City for Science and Technology, Riyadh, Saudi Arabia., Al-Romaih K; From the National Centre of Genomic Technology, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia., Bakheet R; From the National Centre of Genomic Technology, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia., Mahmoud L; From the National Centre of Genomic Technology, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia., Al-Harbi N; From the Biomedical Physics, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia., Alshareef I; From the National Centre of Genomic Technology, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia., Judia SB; From the National Centre of Genomic Technology, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia., Aharbi L; From the National Centre of Genomic Technology, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia., Alzayed A; From the Saudi Human Genome Project, King Abdulaziz City for Science and Technology, Riyadh, Saudi Arabia., Jabaan A; From the Saudi Human Genome Project, King Abdulaziz City for Science and Technology, Riyadh, Saudi Arabia., Alhadrami H; From the Department of of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia., Albarrag A; From the Center of Disease Control, Saudi Arabia Ministry of Health, Riyadh, Saudi Arabia., Azhar EI; From the Department of of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia., Al-Mozaini MA; From the Department of Infection and Immunity, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia.
Jazyk:	angličtina
Zdroj:	Annals of Saudi medicine [Ann Saudi Med] 2020 Sep-Oct; Vol. 40 (5), pp. 373-381. Date of Electronic Publication: 2020 Oct 01.
DOI:	10.5144/0256-4947.2020.373
Abstrakt:	Background: The pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) has prompted a need for mass testing to identify patients with viral infection. The high demand has created a global bottleneck in testing capacity, which prompted us to modify available resources to extract viral RNA and perform reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) to detect SARS-COV-2. Objectives: Report on the use of a DNA extraction kit, after modifications, to extract viral RNA that could then be detected using an FDA-approved SARS-COV-2 RT-qPCR assay. Materials and Methods: Initially, automated RNA extraction was performed using a modified DNA kit on samples from control subjects, a bacteriophage, and an RNA virus. We then verified the automated extraction using the modified kit to detect in-lab propagated SARSCOV-2 titrations using an FDA approved commercial kit (S, N, and ORF1b genes) and an in-house primer-probe based assay (E, RdRp2 and RdRp4 genes). Results: Automated RNA extraction on serial dilutions SARS-COV-2 achieved successful one-step RT-qPCR detection down to 60 copies using the commercial kit assay and less than 30 copies using the in-house primer-probe assay. Moreover, RT-qPCR detection was successful after automated RNA extraction using this modified protocol on 12 patient samples of SARS-COV-2 collected by nasopharyngeal swabs and stored in viral transport media. Conclusions: We demonstrated the capacity of a modified DNA extraction kit for automated viral RNA extraction and detection using a platform that is suitable for mass testing. Limitations: Small patient sample size. Conflict of Interest: None.
Databáze:	MEDLINE
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