Presence of HPV-16 E5 DNA and expression of the specific mRNA in M14 and FRM cells after infection with HPV-16 E5 retroviral vector. The retroviral vector containing HPV-16 E5 gene was obtained by the transfection of Phoenix A retroviral producer cells with the LZRSpBMNZ-E5 plasmid. The control retroviral vector was obtained by the transfection of Phoenix cells with the empty LZRSpBMNZ plasmid. Cells were infected with either recombinant retrovirus or with the control retrovirus. Total DNA or RNA (1 μg) extracted from cells 96 h post infection were reverse transcribed and amplified with E5P65 sense (TGC ATC CAC AAC ATT ACT GGC G) and E5M3AS antisense (AAC ACC TAA ACG CAG AGG CTG C) primers. Upper panel: FRM cells; Lower panel: M14 cells. Lane 1: DNA from cells infected with the control retrovirus; Lane 2: DNA from cells infected with the HPV-16 E5 retrovirus; Lane 3: DNA digested total RNA from cells infected with the HPV-16 E5 retrovirus; Lane 4: Non retrotrascribed DNA digested total RNA from cells infected with the HPV-16 E5 retrovirus; Lane 5: No template negative control; Lane 6 positive control (0.5 μg Siha cell DNA). MW: DNA molecular weight marker VIII (Roche Biochemicals SpA): arrows on the left-hand side indicate the bp length of some reference bands. The band with size of 160 bp (left sided empty arrow) demonstrate the presence of viral E5 sequence and its transcription. Four independent experiments gave similar results.