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. 2010 Nov 11;6(11):e1001183. doi: 10.1371/journal.ppat.1001183

Figure 2. Characterization of virion production and replication of viruses with Myc-tagged p12 proteins.

Figure 2

(A) Western blot analysis of Myc-tagged p12 proteins. Equal amounts of plasmids, coding for 3xMyc and 1xMycR viruses were transfected into 293T cells. Mock are cells transfected with no DNA. Virions were purified from culture supernatants two days posttransfection by ultracentrifugation through 25% sucrose cushions. Tagged-p12 proteins in virion pellets were analyzed by Western blot using anti-Myc monoclonal antibody (9E10). The migration of the cleaved Myc-tagged p12 protein is shown, highlighting the difference in size between the triple and single Myc-tagged p12. (B) Replication kinetics of wt and 1xMycR viruses. Naïve NIH3T3 cells were infected with equivalent amounts of wt or 1xMycR virions (normalized by RT activity) and replication kinetics were determined by radioactive exogenous RT assay. Culture supernatants were harvested at the indicated days postinfection and the arrows indicate the days at which the infected cultures were diluted. Virion level in the sample is directly correlated to the intensity of the radioactive signal [57]. Mock indicates infection with culture medium with no virus.