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Fig. 2 | Virology Journal

Fig. 2

From: Influenza A virus NS1 optimises virus infectivity by enhancing genome packaging in a dsRNA-binding dependent manner

Fig. 2

Effects of NS1 on virus yield. a 293 T RIG-I KO cells were infected with PR8-delNS1 at MOI 0.01 for 1 h and immediately mock transfected or transfected with empty plasmid control (E.C.) or those encoding GFP or NS1 protein. Virus yields were measured 48 h post-infection by plaque assay using MDCK cells. The data shown are the mean ± standard deviation from at least three independent experiments (n = 3). The Student’s t test was used for statistical analysis (n.s. not significant, **P < 0.01). b 293 T RIG-I KO cells were infected as in (a). Cells and culture supernatant were collected to prepare cell lysate and purified virion, respectively. Equal volumes of purified virus were subjected to immunoblot analysis using anti-NP, anti-M1, anti-HA and anti-lamin B antibodies. c 293 T RIG-I KO cells were infected with PR8-delNS1 at MOI 0.1 for 1 h and immediately transfected with NS1 expression plasmid or empty control plasmid. Cells were stained by fluorophore probe targeting segment 5 vRNA (vRNA, Red), anti- NS1 (green), anti-M1 (white), anti-HA (white) and DAPI (blue) 24 h after infection and transfection, and imaged by confocal microscopy. Scale bar, 10 μm. d Supernatant collected from (a) and virus quantity (HAU) was determined by HA assay using 0.5% chicken RBC. e vRNA was extracted from equal HAU from (a) and quantified by RT-qPCR. Levels of viral genome of virus containing supernatant obtained from E.C. transfected cells were set to 1. The Student’s t test was used for statistical analysis (*P < 0.05, **P < 0.01). The data shown are the mean ± standard deviation from at least three independent experiments (n = 3). f Cells in (a) were subjected to extraction for total RNA and vRNA was quantified by qPCR. Levels of viral genome obtained from E.C. transfected cells were set to 1. The Student’s t test was used for statistical analysis (n.s. not significant, *P < 0.05, **P < 0.01). The data shown are the mean ± standard deviation from at least three independent experiments (n = 3)

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