Fig. 3.

(A) Venus RNA analysis following high MOI infection of A549 cells with PLV, EV, IV, or LV reporter viruses. The assay was performed entirely in 96-well format and directly from cell lysates. Infection start times were staggered to allow simultaneous cell lysis. Ct values from Venus RT-qPCR are plotted, starting from Ct 20. “no RT” samples were from the LV infected cell lysates with the RT enzyme omitted from Venus RT-qPCR. (B) Ct values for EV, IV, and LV infections normalized against the Ct value for PLV infection at each time-point. This normalization uncovers promoter-dependent accumulation of Venus RNA (i.e., Venus mRNA). (C) Venus mRNA accumulation at 1, 4, and 8 hpi following EV, IV, or LV infection of A549 cells, in the absence (untreated) or presence of 1 μg/ml AraC. Normalized abundance is plotted. EV data were normalized to 1 hpi EV untreated, IV data were normalized to 4 hpi IV untreated, and LV data were normalized to 8 hpi LV untreated. (D) DNA analysis for LV viral genomes following infection of A549 cells. Shown are normalized qPCR signals for Venus DNA for input (1 hpi) and 8 hpi in the absence or presence of 1 μg/ml AraC.