FIG. 9.

Amplification and packaging of the plasmid pSA1 in Vero cells by ts1249 and ts1208. One set of cell monolayers was transfected with pSA1, and cells subsequently were infected with wt HSV-1, gCB, ts⁺1249MR, or ts1249 or were mock infected (mi) in the presence of cycloheximide at 36.5°C. After 2 h, the cycloheximide block was removed and the samples were transferred to 38.5°C. The second set of cell monolayers was transfected with pSA1, and cells were subsequently infected with ts1208, ts⁺1208MR, or wt HSV-1 or were mock infected. These samples were incubated only at 39.5°C in the absence of cycloheximide. At 20 h p.i., the cells were harvested and total cellular DNA and DNase-resistant DNA were prepared and digested with DpnI and EcoRI (a, c) or BamHI (b, d). Southern blot analysis was carried out, using ³²P-labeled plasmid vector pAT153 (a, c) or the ³²P-labeled, cloned HSV-1 genomic fragment BamHI g (b, d) as a probe. The numbers at the bottom of the lanes indicate the percentage of radioactivity present in the band in the DNase-treated sample relative to that in the band in the total DNA sample. Note that gCB BamHI g is smaller than that of wt HSV-1 strain 17.