High-surety isothermal amplification and detection of SARS-CoV-2, including with crude enzymes

Sanchita Bhadra; Timothy E Riedel; Simren Lakhotia; Nicholas D Tran; Andrew D Ellington

While the NB and Tholoth LAMP-OSD assays were performed using 5 primers (FIP, BIP, F3, B3, . CC-BY-NC-ND 4.0 International license author/funder. It is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.04.13.039941 doi: bioRxiv preprint and LB), the Lamb LAMP-OSD assay was tested using either 5 primers (FIP, BIP, F3, B3, and LB) or 6 primers (FIP, BIP, F3, B3, LB, and LF). Following 90 min of amplification at 65 °C, presence or absence of OSD fluorescence at endpoint was visually observed. As shown in Figure 3 , presence of SARS-CoV-2 genomic RNA resulted in bright, easily detected fluorescence in all three LAMP-OSD assays. The 6-primer version of Lamb LAMP-OSD could detect fewer genomic RNA copies compared to the 5-primer version of the assay. In contrast, all assays showed no signal in the presence of only human genomic DNA.