Figure 3.

Silencing of Sl2/3-MMP expression by RNAi. A, Semiquantitative reverse transcription (RT)-PCR analysis of Sl2-MMP and Sl3-MMP expression in hypocotyls and leaves of 24-d-old tomato wild-type seedlings (WT) and three independent transgenic lines harboring an HP construct for posttranscriptional gene silencing (HP1, HP2, and HP4). After RT, cDNAs of Sl2- and Sl3-MMP and the actin control were detected by 35 and 30 cycles of PCR, respectively. Tomato genomic DNA was used as a control (C). B, Phenotypes and Sl-MMP protein levels of RNAi plants. At top, a 2-week-old tomato plant silenced for Sl-MMP expression is shown on the left, next to a wild-type seedling on the right. Ten plants of the segregating F1 progeny of a primary RNAi transformant were analyzed for the presence of the silencing construct by PCR amplification of the nptII antibiotic resistance marker (middle) and Sl2/3-MMP expression by western-blot analysis (bottom). The seedling phenotype (necrotic hypocotyl) was found to cosegregate with the silencing construct and the loss of Sl2/3-MMP protein expression.