Fig. 1
The design of shRNA-expressing plasmids and antiviral activity. a, Schematic illustration of construction of the dual shRNA expression plasmid (pGPU6/GFP/Neo-shRNA-MN). b, Cells were transfected with shRNA-expressing plasmids for 24 h, transfection efficiency was observed using fluorescence microscope. c, The effect on cell viability was assessed by MTT assay. d, Cytopathic effects caused by PEDV infection. Vero cells were transfected with shRNA-expressing plasmids for 24 h, and then infected with PEDV at 0.1 MOI. At 24 h post-infection, cytopathic effects were observed using microscope. e-g, At 48 h post-infection, cells were harvested and immunoblotted with antibody to viral protein PEDV-N, with actin as a control. N/A, the change in abundance of PEDV-N was analyzed by densitometry analysis using Image-Pro Plus Software and normalized to actin. Under the same experimental conditions, total RNA was isolated and PEDV mRNA was quantified by qRT-PCR normalized against GAPDH (f), and viral yields in the medium were determined by TCID50 (g). These experiments were performed two times with three replicates in each experiment. Significance was determined by one-way analysis. Values represent means and SD for triplicates, *p < 0.05