Fig. 4

Viral ORF7 level was further reduced by transfection with LNA modified AS-ONs in PAMs. Antisense oligonucleotides (YN8, LNA-YN8-A and LNA-YN8-B) were used in the transfection in PAMs with different concentrations. The RNA (a) and protein (b) levels of gene ORF7 were more significantly inhibited by the treatments with LNA modified YN8 than by DNA AS-ON YN8 transfection. After virus challenge and transfection with AS-ONs, total RNA was isolated for RT-qPCR analysis and the total protein was extracted for western blot analysis. The ΔΔCt method for relative quantification of gene expression was used to determine viral ORF7 RNA levels. The Y-axis (a) shows the relative RNA levels of the ORF7 gene for each treatment after normalization to the non-transfected reference sample (PRRSV only). Transfection with 8 μM YN8 or 4 μM LNA-YN8-A or LNA-YN8-B completely blocked the synthesis of viral protein (b).The histogram and blots shown here are representative data from three independent experiments. β-actin was used as internal control in both RT-qPCR and western blot analysis. NS: not significant; *: P < 0.05; **: P < 0.01