Novel SIV-rtTA variants replicate efficiently in primary macaque PBMC. (A) PBMC isolated from cynomolgus macaques were infected with the original or LTR-mutated SIV-rtTA variants. For comparison, cells were infected with SIVmac239. Furthermore, we included the SIV-rtTA-mTat variant in which Tat had been mutated . Cells were infected with an equal amount of virus (corresponding to 10 ng CA-p27) for 16 h, washed and cultured with dox. Replication was monitored by measuring the reverse transcriptase level in the culture supernatant (B) PBMC isolated from rhesus macaques were infected with the indicated SIV-rtTA variants and SIVmac239, using comparable infectious titers (based on titration in TZM-bl cells). Cells were inoculated in the presence of dox and the cultures were split seven days later with half of the cells continuing to receive dox (closed symbols) and the other half receiving no further dox treatment (open symbols). Fresh, uninfected anti-CD3 stimulated cells from allogeneic macaque donors were added every two weeks. Replication was monitored by measuring the viral RNA copy number in the culture supernatant.