To further define what sequence within SL3-2-1 is bound by eEF1Bc, we used complementary DNA oligos to partially convert portions of SL3-2-1 into duplexes (RNA/DNA hybrids) as shown The TCV RdRp assay had two steps: first, the shown components were incubated at room temperature to facilitate their interaction, followed 5 min later the addition of the shown component and the ribonucleotides to start RNA synthesis. The RdRp activity in samples containing the template RNA and the RdRp were chosen as 100% (lanes 5-6 and 9-10). The RNA transcript (20 pmol), eEF1Bc (20 pmol) and purified TCV RdRp (2 pmol) were used in these assays. (C) The effect of eEF1Bc on the TCV RdRp activity with additional templates. One of the templates was SL3-2-1 m(+) with a point mutation within the promoter sequence (carrying SL1m mutation), which is being used more efficiently than the wt SL3-2-1(+) by the TCV RdRp in vitro. The second template was RI-III ( in Figure 3A . EMSA assay with purified recombinant eEF1Bc revealed that the very 39-terminal SL1 region had to be ''free'' (not part of the duplex) for eEF1Bc to bind efficiently to the SL3-2-1 RNA (compare lane 1 with lane 5 in Figure 3A ).