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