Figure 4.

β-Estradiol-Inducible Silencing of pdNAD-MDH in Mature Plant Rosettes.

XVE OLEXA:miR-mdh-1 plants photographed before and after a 6-d treatment with β-estradiol or a mock treatment.

(A) Photographs in the top row were taken before β-estradiol treatment (D0) and those in the bottom row after 6 d (D6). β-Estradiol solution (20 µM) was sprayed every second day onto the entire rosette. β-Estradiol was applied to wild-type (Col) plants as a control. Mock-treated samples were sprayed with water containing the same amount of DMSO as the treatment solution (used to dissolve the β-estradiol). Red arrows indicate examples of white, newly emerging leaves.

(B) Immunoblot detection of pdNAD-MDH in total protein extracts of wild-type and XVE OLEXA:miR-mdh-1 plants before (D0) and 6 d into β-estradiol treatment (D6). For the treated XVE OLEXA:miR-mdh-1 plants, proteins were extracted from the old, green leaves (D6o) and young, white leaves (D6y) separately. Gels were loaded on an equal leaf area basis. The migration of molecular mass markers is indicated on the left. pdNAD-MDH and actin (as a loading control) were detected concurrently on the same membrane using secondary antibodies conjugated to different infrared fluorescence dyes (800CW for pdNAD-MDH and 680RD for actin).

(C) As for (B), but with old leaves from mock-treated samples.