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. 2002 Dec;76(24):12963–12973. doi: 10.1128/JVI.76.24.12963-12973.2002

FIG. 3.

FIG. 3.

Effect of siRNAs on different steps in the HIV-1 life cycle. (A) MAGI cells were transfected with siRNAs directed against either HIV-1 tat (tat1, lane 1; tat2, lane 2; and tat3, lane 3), p65 (lane 4), HTLV-1 tax (lane 5), or Oligofectamine alone (OF, lane 6) for 24 h and were then infected with HIV-1 for another 24 h. Hirt lysates were prepared from these cells, and the cytoplasmic DNA was amplified with oligonucleotide primers to assay for either HIV-1-specific strong-stop (SS) (+43/+65 and +182/+158) (top gel), first-strand jump (−49/−30 and +93/+70) (second gel), full-length (FL) viral DNA (+96/+118 and +234/+212) (third gel), or the mitochondrial cytochrome c oxidase (CO) gene (bottom gel). As controls, cytoplasmic DNA from mock-infected (lane 7) and different amounts of NL4.3 plasmid DNA (20, 2, and 0.2 pg, lanes 8 to 10) were also amplified. (B) Chromosomal DNA was prepared from MAGI cells that were transfected with siRNAs directed against either HIV-1 tat (tat1, lane 1; tat2, lane 2; and tat3, lane 3), p65 (lane 4), HTLV-1 tax (lane 5), or Oligofectamine alone (OF, lane 6) for 24 h and were then infected with HIV-1 for 10 days. Portions of HIV-1 gag (top gel) and cellular α-actin (lower gel) genes were amplified as detailed in Materials and Methods. As controls, 1 ng (lane 7) and 4 ng (lane 8) of NL4.3 plasmid DNA were also amplified.