Figure 1

Characterisation of VSV-G expression plasmids. A) Map of VSV-G expression plasmids. Wild type (wt) or codon optimised (syn) open reading frames of VSV-G are flanked by the human cytomegalovirus immediate early promoter/enhancer region (CMV) and the bovine growth hormone poly(A) signal (pA). Angled black arrows mark the transcriptional start point. The pIG^wt vector contains intron A and flanking untranslated exonic regions E1 and E2 of the cytomegalovirus immediate early gene. In pIΔIG^wt the exon boundaries were precisely fused by deleting the intron. B) Northern blot analysis. Cells were cotransfected with the indicated VSV-G expression plasmids, a codon optimised HIV-1 gag-pol expression plasmid (Hgp^syn) and the lentiviral vector construct VICGΔBH containing a GFP expression cassette. Poly(A) RNA was isolated from transfected cells and analysed by Northern blot with a probe spanning the transcribed region of the BGH poly(A) signal present on all VSV-G transcripts and the positive 5 kb HIV-1 gag-pol transcript. C) Western blot analyses. Cells were cotransfected with the indicated VSV-G expression plasmids, an SIV gag-pol expression plasmid (SgpΔ2) and the lentiviral vector construct VICGΔBH containing a GFP expression cassette. Monoclonal antibodies to HIV-1 p24 capsid protein, which is cross reactive to SIV p27, or to VSV-G, respectively, were used for detection of the viral proteins in lysates of transfected cells.