Journal Article

LuNER: Multiplexed SARS-CoV-2 detection in clinical swab and wastewater samples

Fan, Vinson B., Gildea, Holly K., Stahl, Elizabeth C., Giannikopoulos, Petros, Hamilton, Jennifer R., Sylvain, Iman A., Metzger, Matthew, Keller, Amanda, Doudna, Jennifer A., Tsuchida, Connor A., McDevitt, Shana L., Nelson, Kara L., Pestal, Kathleen, Ringeisen, Bradley R., McElroy, Matthew, Urnov, Fyodor D., Hirsh, Ariana, Ehrenberg, Alexander J., Gopez, Allan R., Tsui, C. Kimberly, Ciling, Alison, Kantor, Rose, Witkowsky, Lea B., Lin-Shiao, Enrique, Moehle, Erica A., Williams, Clara

2021

Overview

Clinical and surveillance testing for the SARS-CoV-2 virus relies overwhelmingly on RT-qPCR-based diagnostics, yet several popular assays require 2–3 separate reactions or rely on detection of a single viral target, which adds significant time, cost, and risk of false-negative results. Furthermore, multiplexed RT-qPCR tests that detect at least two SARS-CoV-2 genes in a single reaction are typically not affordable for large scale clinical surveillance or adaptable to multiple PCR machines and plate layouts. We developed a RT-qPCR assay using the Luna Probe Universal One-Step RT-qPCR master mix with publicly available primers and probes to detect SARS-CoV-2 N gene, E gene, and human RNase P (LuNER) to address these shortcomings and meet the testing demands of a university campus and the local community. This cost-effective test is compatible with BioRad or Applied Biosystems qPCR machines, in 96 and 384-well formats, with or without sample pooling, and has a detection sensitivity suitable for both clinical reporting and wastewater surveillance efforts.

• Publisher: Public Library of Science,Public Library of Science (PLoS)
• Subject: Analysis / Biology and life sciences / Consortia / Coronaviruses / COVID-19 / COVID-19 - virology / Diagnostic tests / DNA Primers - genetics / E protein / Environmental engineering / Funding / Genes / Genomics / Human subjects / Humans / Medical laboratories / Medicine and Health Sciences / Multiplexing / N gene / Public health / Real-Time Polymerase Chain Reaction - methods / Research and analysis methods / Ribonuclease / Ribonuclease P / Ribonuclease P - genetics / RNA, Viral - genetics / SARS-CoV-2 - genetics / Sensitivity and Specificity / Severe acute respiratory syndrome / Severe acute respiratory syndrome coronavirus 2 / Specimen Handling - methods / Testing / Viral diseases / Viruses / Wastewater / Wastewater - virology / Wastewater-Based Epidemiological Monitoring