Fig 1.

pUL51 is required for cleavage of HCMV genome concatemers, but not for viral DNA replication. (A, left) DNA replication of the HCMV-UL51-ddFKBP mutant in the presence or absence of shield-1. HFF infected with the mutant were harvested on days 1 to 6 p.i. Total DNA was isolated and applied to slot blot hybridization with a probe specific for the b-repeat region. (Right) Signals were quantified using a phosphorimager. (B) Detection of unit-length genomes by pulsed-field gel electrophoresis. Cells infected with the HCMV-UL51-ddFKBP mutant were kept in the presence or absence of shield-1 for 5 days, and total DNA was subjected to pulsed-field gel electrophoresis followed by hybridization to the HCMV-specific probe. (C) Detection of free genomic ends. (Left) HCMV-UL51-ddFKBP-infected HFF cultivated with or without shield-1 were harvested on day 5 p.i. Total DNA was extracted, cut with HpaI, and analyzed by Southern blotting using the b-specific probe. (Right) Schematic drawing of the four isomeric forms of the HCMV genome, showing the orientation of the unique long (UL) and unique short (US) regions (arrows) and the fragments detected by the probe, with their sizes given below the diagrams.