Analysis of integrated HIV-1 vector DNA in primary human lymphocytes.

(A) FACS analysis of EGFP expression in mitogen-stimulated and resting primary human PBLs transduced with pHR′PGK-EGFP vector in the presence (+) or the absence (Δ) of HIV-1 accessory proteins (APs) at 5 days after transduction. (B) Detection of integrated HIV-1 provirus by using Alu-HIV-1 LTR PCR. Genomic DNA isolated from mitogen-stimulated and resting primary human PBLs, untransduced (UT) or transduced with pHR′PGK-EGFP vector in the presence (+) or the absence (Δ) of HIV-1 accessory proteins (APs) at 5 days after transduction shown in Figure5A was subjected to Alu-HIV-1 LTR PCR procedure (PCR-1 + PCR-2) or only to the second round of amplification (PCR-2 alone). The amplification products were resolved in an agarose gel and bands were detected by Southern blot analysis by using an LTR-specific probe.