Figure 4

Mapping of HPV11 TSSs and splice junctions in U2OS cells via 5′ RACE. U2OS cells were transfected as follows: 500 ng of the HPV11wt genome (lane 1), 500 ng of the HPV11wt genome together with 2 μg of the linearized pBabe-Neo construct (lanes 2 and 3), 500 ng of the HPV11E8- genome (lane 4), or mock transfection (lane 5). PolyA⁺ mRNA was extracted at 4 and 10 days post-transfection, and 5′ RACE analysis was performed using the HPV11-specific primers pr855c (A), pr1265c (B), pr1558c (C), pr2891c (D) and pr3692c (E). RT-PCR analysis was performed using the HPV11-specific primers pr1241 and pr3692c (F). All distinct PCR products (marked with letters from A to Q; the same designations are used in the map represented in Figure 5) were cloned and sequenced for the analysis of TSSs and/or splice junctions.