FIG. 5.

An inverted distal enhancer augments viral DNA replication. (A) Schematic diagram of WT and r-579/-300. Base positions of PstI, NdeI, and NcoI sites and probe (solid bar) coordinates are given relative to the RNA start site of the MIE promoter. INV, inverted distal enhancer. (B) Southern blot analyses of PstI and combined NdeI and NcoI RFLPs of WT, r-579/-300, and rΔMSVgpt. Predicted sizes of RFLPs are shown. (C) Abundances of WT, r-579/-300, and rΔ-300/-579 genomes in HFF cells at an MOI of 1.0 on days 2 and 3 p.i. (DPI 2 and 3). (D) DNA replication rates of WT, r-579/-300, and rΔ-300/-579 genomes in HFF cells at an MOI of 0.005 on days 6 and 7 p.i. Infections for which results are shown in panels C and D were performed in parallel by methods described in the legend to Fig. 1C. (E) Abundances of WT, r-579/-300a, r-579/-300b, and rΔ-300/-579 genomes in HFF cells at an MOI of 0.005 on day 7 p.i. Viruses r-579/-300a and r-579/-300b were derived from independent transfection-recombination procedures. The abundances of viral genomes at 4 h, 2 days, and 3 days p.i. at an MOI of 1 were equivalent for all four viruses (data not shown). Analysis was performed as described in the legend to Fig. 1C.