Figure 4: Replication of full-length STMV RNA bearing mutations in the N-terminal 13aa motif.

(A) Mutations engineered into the N-terminal 13aa motif of CP. Positively charged amino acid residues are indicated by a + sign. Alanine or lysine substitutions at designated positions are underlined. Nomenclature used to designate each mutation is the same as described under the Fig. 3 legend. (B–E) Progeny analysis. Total RNA and protein samples were isolated from leaves harvested at 4 dpi. Panels B and D summarize Northern blot analysis of (+) and (−)-strand accumulation for the indicated variants and RT-PCR analysis. Panel C summarizes the absolute values of accumulated (+) and (−)-strand progeny RNA. (E) Western blot analysis of CP expression for indicated variants. Following co-infiltration of pRP and each variant agrotransformant progeny were analyzed by Northern blot hybridization, RT-PCR, Western blot analysis and the absolute values of accumulated (+) and (−)-strand progeny RNA.as described in the Figs 2 and 3 legends. (F) Detection of CP 3A by Western blot analysis using a 10-fold excess of the indicated protein samples. Positions of STMV and STMVΔ150 are indicated to the left in panels B and D. Ribosomal RNA (rRNA) represents loading controls for Northern blots. Position of monomeric (1×) and dimeric (2×) forms of STMV CP is indicated to the right in panel E. Accumulation of (+) and (−)-strand progeny was quantitated as described in the Fig. 2 legend.