FIG. 4.

Wild-type and mutant MVMp virus replication in a single-round infection. A9 cultures were infected with 5 CFU of primary stocks of either wild-type (wt) or mutant (585A/93G or 588A/96S) MVMp virus/cell and further incubated in culture medium supplemented with neuraminidase (from 6 h p.i. on) to prevent new rounds of infection. Cells were harvested at the indicated times p.i. and processed for the analysis of virus replication. (A) Southern blotting analysis of viral DNA intermediates. mRF, monomer RF; dRF, dimer RF; ssDNA, single-stranded genomic DNA. (B) Western blotting analysis of viral nonstructural protein production. The brackets encompass the phosphorylated and un(der)phosphorylated forms of the respective polypeptides. (C) Titration of progeny viruses by replication center assays on A9 indicator cell monolayers. Lane 1, amount of progeny viruses in adherent A9 monolayers; lane 2, total amount of viral particles present in adherent and detached cells, as well as in the culture medium; nd, not determined.