FIG. 2.

siRNA knockdown of TNF-α in MDM has no effect on DV replication. MDM were transfected with individual TNF-α siRNAs by two rounds of 4-h transfection with 1.6 μM TNF-α siRNA 4, 5, or 6 infected with DV at an MOI of 5, and at 32 h p.i. cell culture supernatants were analyzed. (A) TNF-α production (ELISA). Results were expressed as a percentage of TNF-α released from DV-infected mock-transfected MDM (2,027 pg/ml/24 h). Values are means ± standard errors of the means of two wells. (B) DV replication determined by infectious virus release (PFU/ml, black bars) and negative-strand RNA levels (white bars) in infected cells. In vitro-transcribed copy number standards from DV2 capsid protein were used to quantitate negative viral RNA using a tagged RT real-time PCR. Values represent means ± standard errors of the means from duplicate samples in RT real-time PCR of a representative experiment per ng cyclophilin RNA. Experiments were replicated (n = 2).