The TCV RdRp reactions were carried out as previously described for 2 h at 25uC [36] , except using 7 pmol template RNA and 2 pmol affinity-purified MBP-p88C. Different concentrations of eEF1Bc (6xHis-affinity purified recombinant Tef3p obtained from E. coli or Flag-affinity purified HF-Tef4p obtained from yeast) were added to the reaction at the beginning or as indicated in the text and Figure 2 . legend. The 32 P-labeled RNA products were analyzed by electrophoresis in a 5% PAGE/8 M urea gel [63] . The 86-nt 39 noncoding region of TBSV genomic RNA and its mutants were used as the template in the RdRp assay [24, 36] . RNA templates were generated with T7 transcription using PCR products obtained with the following primers: #1662 (TAATACGACTCACTATAG GACACG GTTG ATC TC ACC-CTTC) and #1190 (GGGCTGCATTTCTGCAATG) for SL3-2-1(+), #1662 and #4390 (GGGCTGCACAAGTGCAAT-GTTCCGGTTGTCCGGT) for SL3-2-1cuug(+). SL3-2-1m(+) RNA was generated with T7 transcription on PCR products amplified with primers #1662 and #1190, on a plasmid template harboring GGGCU nucleotide-deletion in SL3 region as described [39] . A duplex RNA was generated by hybridizing SL3-2-1(+) and SL3-2-ds1(-) made by T7 transcription of the PCR product using primers #4361 (GTAATACGACTCACTA-TAGGGCTACTTCCGGTTGTCCGGTAGTGCTTCC) and 