Fig. 7

LlMYB305 binds the promoter of LlC3H18 and activates its expression. (A) Diagram of the LlC3H18 promoter. The W-box elements are marked with blue triangles. The truncated fragments used for the yeast one-hybrid (Y1H) assay are marked with black lines. The mutant fragment used for the Y1H assay is marked with a red line. (B) A Y1H assay for LlMYB305 and the promoter of LlC3H18. Fragment activity was analyzed by a color change on Ura-/Trp-deficient SD medium following the addition of x-gal. One representative image based on three replicates. (C and D) An electrophoretic mobility shift assay (EMSA) of GST-LlMYB305 and the potential elements from the LlC3H18 promoter. The probe 1 and 2 came from the core sequence of P4 and P5 fragments, respectively. One representative image based on three replicates. (E) Constructs used in the dual-luciferase reporter assay. (F) Detection of the LUC signal in tobacco leaves. One representative image based on three replicates. Scale bar = 1 cm. (G) Measurement of LUC intensity in the dual-luciferase reporter assay. Data are presented as means ± SD of three replicates (Student’s t-test, * P < 0.05). (H-J) Detection of the expression level of LlMYB305 (H), LlC3H18 (I), and LlWRKY33 (J) in LlMYB305-overexpressed lily petals. Data are presented as the mean ± SD of three replicates (Student’s t-test, * P < 0.05). (K-M) Detection of the expression level of LlMYB305 (K), LlC3H18 (L), and LlWRKY33 (M) in LlMYB305-silencing lily petals. Data are presented as the mean ± SD of three replicates (Student’s t-test, * P < 0.05). (N) Phenotypes of lily petal discs under room temperature conditions (RT, 22 °C) and after exposure to heat stress (HS, 40 °C, 12 h). Representative image came from three experiments. Scale bar = 1 cm. (O) Relative ion leakage (%) of discs at 22 °C (RT) and after HS (40 °C, 12 h). Data are presented as the mean ± SD of three replicates (Student’s t-test, * P < 0.05; ND, no significant difference; the SK-LlMYB305 was compared with the SK-II control under the RT or HS condition, respectively. (P) Phenotypes of lily petal discs at RT (22 °C) and after HS (40 °C, 12 h). Representative image based on three experiments. Scale bar = 1 cm. (Q) Relative ion leakage (%) of discs at RT and after HS (40 °C, 12 h). Data are presented as the mean ± SD of three replicates (Student’s t-test, *P < 0.05; ND, no significant difference; TRV-LlMYB305 was compared with the TRV-control under the RT or HS condition, respectively)