Journal: bioRxiv

Article Title: Loss of nsp14-exonuclease activity impairs the replication, proofreading, fitness and pathogenesis of SARS-CoV-2

doi: 10.64898/2026.01.12.698941

Figure Lengend Snippet: VTA cells were infected with nanoluciferase-expressing SARS-CoV-2 WT or AA at 37°C at an MOI of 0.1 for 1hr after which input was removed, monolayers were washed and then exposed to a dose response of 5-fluorouracil (5-FU), β-d-N4-Hydroxycytidine (NHC, EIDD-1931) or GS-441524 in Infection Media. Concurrently, non-infected cells were treated similarly to determine cytotoxicity. After 24hr, viral replication was assessed by NanoGlo Luciferase Assay System (Promega) and cytotoxicity was determined by CellTiterGlo Assay (Promega). Each condition was evaluated in triplicate in two independent studies. Values were normalized to the uninfected and infected vehicle DMSO controls (0 and 100% infection, respectively). Data were fit using a four-parameter nonlinear regression analysis using GraphPad Prism. EC50 and CC50 (cytotoxic concentration at which 50% of cells are viable) values were then determined as the concentration reducing the signal by 50%.

Article Snippet: After incubation, the virus was removed, the monolayers were washed with 100μL of Infection Media, and the cells were treated with three-fold serial dilution series of 5-fluorouracil (5-FU, Sigma), β-d-N4-Hydroxycytidine (NHC, EIDD-1931, MedChemExpress) or GS-441524 (MedChemExpress) in Infection Media, starting at concentrations of 400μM, 20μM, and 20μM respectively.

Techniques: Infection, Expressing, Luciferase, Concentration Assay