Visualization of Early RNA Replication Kinetics of SARS-CoV-2 by Using Single Molecule RNA-FISH Combined with Immunofluorescence.

ABSTRACT: SARS-CoV-2 infection remains a global burden. Despite intensive research, the mechanism and dynamics of early viral replication are not completely understood, such as the kinetics of the formation of genomic RNA (gRNA), sub-genomic RNA (sgRNA), and replication centers/organelles (ROs). We employed single-molecule RNA-fluorescence in situ hybridization (smRNA-FISH) to simultaneously detect viral gRNA and sgRNA and immunofluorescence to detect nsp3 protein, a marker for the formation of RO, and carried out a time-course analysis. We found that single molecules of gRNA are visible within the cytoplasm at 30 min post infection (p.i.). Starting from 2 h p.i., most of the viral RNA existed in clusters/speckles, some of which were surrounded by single molecules of sgRNA. These speckles associated

SUBMITTER: Pathak R 

PROVIDER: S-EPMC10893374 | biostudies-literature | 2024 Feb

REPOSITORIES: biostudies-literature