FIG 2 .

Characterization of Brts105 growth. Viruses were inoculated at a multiplicity of 10 PFU/cell on primary MEFs (mouse embryo fibroblasts) at 33°C for 1 h and then incubated at either 33°C (A) or 40°C (B) for the duration of the experiment. Viral growth was measured by plaque assay at 33°C. The average of three replicates is shown. Dotted lines indicate the lower limit of detection for the plaque assay. (C) Northern blots show the effect of temperature on viral RNA production by the wild type and Brts105. Bands corresponding to viral subgenomic RNAs 4 to 7 are indicated. (D to F) The relative severity of ts phenotypes was characterized by a growth stop/resume assay. Growth was stopped 4.5 h after inoculation by incubating ts virus at 40°C or by treating wild-type virus with 10 µg/ml cycloheximide (CHX). After 3 h of arrested growth, the inhibition was either maintained to the end of the experiment (Stopped) or released (Resumed). Virus growth was measured by plaque assay, and the results for Brts105 are shown in panel E. The averages for 6 replicates are shown. The titer ratio (resumed to stopped) from panel E is shown in comparison to other mutants and CHX treatment in panel F. Statistically significant increases in titer after resumption are indicated (*, P ≤ 0.05; ns, not significant).