Modification of the cytoplasmic tails of BaEV glycoproteins allows efficient pseudotyping of lentiviral vectors. (A) Schematic representation of the WT (BaEVwt) and mutant BaEV (BaEVTR, BaEVRless), RDTR- and MLV-A gps. The 17 aa long cytoplasmic domain of the cat and baboon retroviral gps, RD114 and BaEVwt, respectively, was exchanged for one of the MLV-A glycoproteins resulting in the chimeric RDTR- and BaEVTR-gps, respectively. The R peptide of the cytoplasmic tail of BaEVwt was deleted, resulting in the BaEVRLess mutant gps. Amino acid sequences and different domains for these gps are shown in supplemental Figure 1. (B) Titers of the different pseudotyped LVs encoding GFP, obtained by infection of HEK293T cells with serial dilutions of fresh or 100-fold concentrated vector preparations (infectious units/mL). The percentage of GFP⁺ cells was determined 3 days after infection by FACS (means ± standard deviation [SD], n = 8; ***P < .005). (C) Immunoblots of LV particles displaying the different BaEV-gps at their surface. LVs were purified over a sucrose cushion by ultracentrifugation. The upper part shows staining with antibodies against the surface domain of BaEVwt. The corresponding HIV-1 capsid was revealed to verify equal loading. The positions of the BaEVgps and the HIV-1 capsid (HIV1-CA) are indicated. (D) Pictures show 293T cells 48 hours after transfection with BaEVTR- and BaEVRless-gps.