Transient expression of V5-tagged PTI-1 in vivo . NIH 3 T3 cells were transiently transfected with the following constructs: pcDNA3.1-PTI-1-ORF1-V5 (lane 1), pcDNA3.1-PTI-1-FL-V5 (lane 2), pcDNA3.1-PTI-1-FL-ΔATG-V5 (lane 3) or an empty pcDNA3.1 vector (lane 4). Cells were harvested and lysed approximately 48 hours post transfection. 40 μg of total protein lysate was loaded in all lanes. Expression was analyzed by Western blotting using an anti-V5-antibody. The presence of the V5 tag increases the molecular weight of the expression product by 2.6 kDa. Arrows indicate the bands with potential correspondance to protein products resulting upon translation of ORFs 1, 2, 3 and 4 shown in Figure 1. A background band of unknown origin appears above the ORF3 band in all lanes and serves as a loading control.