Figure 7

Analyses of the effect of the mutated pCD163 on PRRSV-2 invasion. A, B The binding, entry and infection assays with mutated pCD163 for PRRSV-2 strain BJ-4 and HN07-1, respectively. WT or mutated pCD163 constructs were transfected into PK-15 cells. After 24 h, the transfected PK-15 cells were inoculated with PRRSV strain BJ-4 or HN07-1 at a MOI of 1 at 4 °C for 1 h, and then relative quantitation of viral RNA abundance was carried out. For PRRSV entry assay, the unbound viruses were washed away and the inoculated cells were then cultured at 37 °C for 3 h to allow viral entry. The entering viral RNA was analyzed by RT-qPCR. For PRRSV infection, PRRSV strain BJ-4 or HN07-1 was inoculated in the transfected PK-15 cells as described above. PRRSV RNA abundance was tested by RT-qPCR. For the RT-qPCR, PRRSV ORF7 gene was normalized with GAPDH mRNA and relatively quantified by the 2^−ΔΔCT method. In parallel, we have transfected empty vector and inoculated PRRSV as negative control. However, the C(t) value of the negative control was comparable to that of water, which was not included in the manuscript. Data represent means ± SEM of three independent experiments. *p < 0.05, **p < 0.01.