FIG. 4.
HIV-1 reverse transcription in BMVECs proceeds to LTR/U5. (A) Replicate BMVEC cultures (400,000 cells in 60-mm-diameter tissue culture dishes) were infected with 106 RNA copies of HIV-1LAI (100 pg of p24 antigen) for 2 h at 37°C and washed four times with PBS. The cells were harvested using 0.25% trypsin-1 mM EDTA at the indicated times p.i. The cell lysates were subjected to PCR amplification using the primers from the LTR/U5 region, the pol gene, or 2LTR circles. The hybridization was positive only with the LTR/U5 probe; the hybridizations with the probe for pol or 2LTR circles were negative. (B) Dilutions of PCR lysates prepared from 8E5/LAI cells containing one HIV-1 provirus per cell were amplified with the same set of primers used for panel A. (C) Cell lysates from CEM lymphoblastoid cells or BMVECs infected with HIV-1 for 9 days were subjected to PCR amplification using the primers from the pol gene.