(A) Recombinant virus was produced in 293T cells transfected with pTM-pol I-WSN-All and expression vectors for NP, PB1, PA, PB2, and WT or K627E PB2-TAP. PB2 and PB2-TAP were detected by western blot of cell lysate or cell-free virions with anti-PB2 serum. (B) PB2-TAP was precipitated with rabbit IgG-agarose from virus-producing cell lysates. Co-precipitating NP was detected by anti-NP western blot. (C) Purified virus was lysed and separated on a linear 30–60% glycerol gradient. Where indicated, 1 % deoxycholate (DOC) was present during viral lysis. Fractions were collected and PB2 and NP were detected by western blot. Non-specific bands at the top of the gradient are indicated (*).