FIG. 6.

N- and C-terminal chimeras of Nef. PCR was used to fuse the N terminus of HIV-1 Nef to the C terminus of HIV-2 or SIV Nef. Likewise, the N terminus of HIV-2 or SIV Nef was fused to the C terminus of HIV-1 Nef. (A) Schematic of the PCR procedure. The procedure takes advantage of the fact that the PPT sequence is highly conserved in each virus, allowing the same internal primers to be used to create each clone. (B) P4 MAGI cell assays were done to test the infectivity of each chimera. Virions used for the infection were normalized by p24 content, and particles were produced in the presence and absence of CsA (10 μM). Each measurement was done in triplicate, and the results shown are from two independent experiments. (C) Immunoblot analysis was performed on virions to determine the relative amount of each chimeric Nef present. Loading was normalized by p24 antigen concentration.