JoVE Encyclopedia of Experiments
Microbiology
0 views • 3:35 min • September 30th, 2026
Take mammalian cell cultures. Infect the test culture with a virus, leaving the control uninfected.
Allow the virus adsorption, then replace the medium.
The virus enters the cell and releases its genome. The virus expresses a virulence protein that destabilizes host transcription factor complexes and suppresses host transcription.
The control cells, however, maintain active transcription.
Add a nucleoside analog that gets incorporated into nascent RNAs.
Harvest the washed cells.
Fix the cells to preserve their structure. Centrifuge the cells and wash them with a buffer.
Add a detergent to permeabilize the cells, then wash.
Add a chemically modified fluorescent dye that reacts with the nucleoside analog to label nascent RNAs; then wash.
Perform immunostaining to fluorescently label viral antigens, marking infected cells.
Use flow cytometry to detect host RNA and viral antigen fluorescence.
Compare the RNA fluorescence intensity between the control and test populations. A downward shift in the test population, indicating reduced RNA fluorescence, suggests virus-induced transcriptional suppression.
As per experimental design, infect the 293 cells with MP-12 at an MOI of three. Remove the growth m
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