Fig 2.

Venus/M093 fusion protein is incorporated into MYXV virions. (A) Temporal synthesis of Venus/M093 protein. BSC-40 cells were mock infected or infected with vMyx-Venus/M093 at an MOI of 10.0 in the absence or presence of cytosine arabinoside (AraC), indicated by +AraC, for 1 h at 37°C. After adsorption, the inoculum was removed and cells were washed. At the indicated time points, cells were harvested and lysed in radioimmunoprecipitation assay buffer. Clarified cell lysates were subjected to SDS-PAGE, and Western blotting was performed sequentially using the indicated antibodies, followed by an appropriate horseradish peroxidase-conjugated secondary antibody. (B) Venus/M093 fusion protein is incorporated into MYXV IMVs. IMVs purified through a 36% sucrose cushion were treated with NP-40 lysis buffer in the presence or absence of DTT. The detergent-insoluble fraction (I) was separated from the detergent-soluble fraction (S) or left unfractionated as total (T) input protein. The fractions were subjected to SDS-PAGE for Western blot analyses using the antibodies indicated in panel A.