Figure 4.

Transgenic wheat lines L65 and L91 producing fungal gene-derived siRNAs induce silencing of the target mRNA and confer resistance to Pst infection. A, Phenotypes of transgenic lines L65, L91, and control in T2 and T₃ generations. The second leaves of seedlings were inoculated with urediospores of CYR32 and photographed at 16 dpi with Pst in each generation. Control is the transgenic lines carrying empty vector; L65 and L91 are the transgenic lines carrying RNAi constructs. B, Northern-blot analysis of siRNA isolated from T₃ transgenic wheat lines detected with specific probe derived from the PsFUZ7 fragment. U6 is the small nuclear RNA as a loading control. C, Relative expression of PsFUZ7 at 3, 10, and 16 dpi with CYR32 of T₃ transgenic wheat lines L65, L91, and control. Data were normalized to PsEF-1, and the CK-10 d control was set to 1. D, Host response and infection types in T₃ transgenic wheat lines L65, L91, and control assessed at 16 and 25 dpi with CYR32. E, Fungal biomass measured using real-time PCR of total DNA extracted from wheat leaves infected with CYR32 at 16 dpi. Ratio of total fungal DNA to total wheat DNA was assessed using the wheat gene TaEF-1 and the Pst gene PsEF-1. Differences were assessed using Student’s t tests. Values represent the means ± se of three independent samples. Double asterisks indicate P < 0.01. CK, Control.