To analyse the proteins produced during À1 PRF, lacZ-A/ Am/B/Bm/C/Cm-Fluc fragments were cloned inframe with the N-terminal 6xhistidine-tag in pYES2/NT KpnI and XhoI sites (Invitrogen). Reporter fusions were amplified by PCR using pAC-A/Am, pAC-B/Bm or pAC-C/Cm as templates. The resulting plasmids were named pYES2/NT-A/ Am, B/Bm and C/Cm. Protein expression was regulated from GAL1 promoter. Two CfMV encoded proteins, P27 (C-terminal end of ORF2A) and replicase (ORF2B), were cloned into pYES2 (Invitrogen). Translation initiation codons were introduced within the oligonucleotides during PCR. The resulting plasmids were named pYES-P27 and pYES-Rep. Control plasmids, which lacked the translation initiation codons were prepared by PCR-based mutagenesis (pYES-P27DAUG and pYES-RepDAUG) and the resulting plasmids were verified by sequencing. Plasmids encoded auxotrophic marker for uracil (URA3).