Figure 3.

Effect of the fs mutation in the presence and absence of the uORF start codon. (A–E) Transient expression studies of the ANAC082 (A), CIPK6 (B), At3g15430 (C), At5g27920 (D) and OTLD1 (E) CPuORFs. The 35S::UTR:RLUC reporter plasmids carrying the WT CPuORF, the mutant CPuORF lacking the start codon (ΔAUG), the fs mutant CPuORF or fs ΔAUG double mutant CPuORF of each gene was co-transfected with the 35S::FLUC internal control plasmid into MM2d protoplasts by electroporation, and the reporter activities were analyzed as in Figure 2. Means ± S.D. of at least three biological replicates are shown. Each graph is representative of two or more separate experiments using independently prepared protoplasts. Double asterisks indicate a significant difference between two constructs (P < 0.01 by t-test), whereas ‘n/s’ indicates non-significant difference (P ≥ 0.05 by t-test).