Construction and expression of Vpr and C81. (A) Plasmids containing cDNA for Vpr and C81 were generated from HIV-1NL4-3. Schematic presentation of the predicted first α-helical domain (αH1), second α-helical domain (αH2), third α-helical domain (αH3) and the arginine rich region of Vpr are indicated. (B) Western blotting of wild-type Vpr and C81. HeLa, HepG2, and 293 T cells were transfected with pME18Neo encoding Flag-tagged wild-type Vpr or C81, or the empty vector control pME18Neo-Flag, together with pSV-β-galactosidase. Transfected cells were collected and lysed 48 h after transfection. Lysates with equal β-galactosidase activity were subjected to Western blot analysis using anti-Flag.