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. 2020 May 14;9(5):376. doi: 10.3390/pathogens9050376

Figure 4.

Figure 4

ORF3 increased the activation of IKBKB-meditated NF-κB promoter activity. (A) HEK293T cells were grown in 24-well plates and transfected with plasmids expressing ORF3, IKBKB, firefly luciferase driven NF-ҡB promoter and Renilla luciferase. At 24 hpt, cells were treated with 50 ng/mL TNF-α for 8 hrs. Cells were then lysed, and the firefly luciferase and Renilla luciferase activities were measured. Data (n = 3) were shown as the relative firefly luciferase activities normalized to the Renilla luciferase activities. *, p < 0.05. (B) LLC-PK1 cells were transfected with the plasmid expressing IKBKB or/and ORF3 for 24 h following with TNF-α treatment (50 ng/mL) for 8 hrs. Cells were lysed and subjected to total RNA extraction. The RT-qPCR was performed to detect the relative expression of cytokine mRNA. Relative mRNA expression (n = 3) of cytokine mRNA was normalized with the internal control β-actin. Error bars indicated mean± standard error of means of relative normalized mRNA expression from duplicate experiments. *, p < 0.05; ***, p < 0.001.