FIG. 5.

Reiterative primer extension analysis of Ty1 replication intermediates. 5′-end-labeled primers complementary to −sssDNA or +sssDNA were annealed to VLP-derived nucleic acids and repeatedly extended with Taq polymerase. The asterisk at the 5′ end of the primer represents the ³²P label. The sequences at the 5′ ends of the −sssDNA and +sssDNA are indicated. The extension products were loaded on an 8% denaturing polyacrylamide gel along with a sequence which was used as a size marker. The sequence was determined with the primer used in the primer extension assay so that the extension product would migrate like the sequencing fragments bearing the last nucleotide of the strong-stop DNA (indicated by an asterisk in the sequencing gel). In fact, the primer extension product migrated slightly faster, because the primer used in the primer extension assay was phosphorylated at its 5′ end, whereas the primer used in the sequencing reaction was not phosphorylated. PBS, primer binding site.