Figure 7.

An artificial stem-loop structure replacing mSL2 in MYB33 promotes silencing. A, Synonymous mutations were introduced into the MYB33 gene to generate MYB33-mSL2-2 (purple). B, Predicted RNA secondary structure of MYB33 and MYB33-mSL2-2. Structures were predicted via the Vienna RNAfold web server. The heat map indicates the probability of second structure formation, from low (purple) to high (red). The miR159 binding site is outlined in black. The altered mSL2-2 structure is outlined in purple. The mutated nucleotides in mSL2-2 as shown in Figure 7A, are indicated with arrows. C, Number of primary transformants falling into each phenotypic category, as a percentage of the total number of transformants analyzed for each construct (n). D, qRT-PCR measurement of MYB33 and CP1 mRNA levels in primary transformants of MYB33, MYB33-mSL2-2, and mMYB33-mSL2-2. myb33 plants were used as controls. All measurements are relative to CYCLOPHILIN. RNA was extracted from two independent biological replicates, each being composed of rosette tissue sampled from 10 to 15 primary transformants. The two measurements are shown as dots, with the bar representing the mean.