FIG. 9.

In vivo labeling of replicating full-length HCV RNA during cell cycle arrest and release. Huh7 2-3 cells were arrested in G₁ by treatment with aphidicolin for 24 h and subsequently released from the cell cycle block. At 4 h before being harvested, the cells were starved in phosphate-free medium for 1 h, followed by the addition of [³²P]orthophosphate at a concentration of 500 μCi/ml. Total RNA was extracted and analyzed by electrophoresis, and radioactively labeled, newly synthesized HCV RNA was quantitated by densitometry. There was a reproducible increase in HCV RNA synthesis as the cells progressed through S phase, with HCV RNA synthesis peaking approximately 8 h after release from the G₁ block.