CpG enrichment in segment 5 of the IAV genome reduces NP transcript and protein abundance in cells. (A) To assay transcription efficiency, in vitro transcription assays using T7 RNA polymerase were performed using the segment 5 plasmids, with RNA quantified by Qubit at 10, 20, 45, and 90 min. (B) To assay combined transcription and translation in a cell-free system, limiting dilutions of segment 5 plasmids were used in rabbit reticulocyte lysate coupled transcription and translation assays, and NP protein was detected by western blotting using an NP-specific antibody. (C, D) A549 cells were infected at MOI 10 for 10 h, then RNA abundance was examined by northern blotting (C) and protein abundance by western blotting for the indicated species (D).
