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. 2016 Jun 22;6:28481. doi: 10.1038/srep28481

Figure 6. NS1 and NS1-70 block the association of p65 with the κB DNA element.

Figure 6

293T cells were transfected with NS1-HA or NS1-70-HA expression plasmid or empty vectors for 30 h, The cells were mock-treated or treated with TNF-α (10 ng/ml) for 60 min. Nuclear extracts were then isolated and subjected to DNA affinity-binding assay by using Streptavidin magnetic particles and biotinylated dsDNA oligonucleotides containing the κB DNA element sequence or the control oligonucleotide YY1 promoter sequence. DNA-bound proteins and 5% of input nuclear extracts were analyzed by immunoblotting to detect p65 using anti-p65 antibody and NS1-HA together with NS1-70-HA using the anti-HA antibody. HDAC1 was used as a loading control to demonstrate that equal amounts of nuclear proteins were present in various samples. One of three experiments is shown.