Fig. 1. Expression patterns and phenotypic analysis of OsGZ7. A, Expression of OsGZ7 in various tissues under different zinc (Zn) levels [0.04, 0.4 (control), and 40 μmol/L]. Thirteen-day-old seedlings were transferred to 1/2 Kimura’s B solution with different Zn concentrations for 3 d before sample collection for RNA extraction and expression analysis. OsActin was used as the internal control. B, β-glucuronidase (GUS) expression of OsGZ7. Thirteen-day-old seedlings were hydroponically grown for 7 d under Zn-normal (0.4 μmol/L) and Zn-deficient (0.04 μmol/L) conditions. Blue indicates GUS activity in root (a), root tip (b), root hair (c), basal stem (d), and leaf (e). In (b)-(e), scale bars, 2 mm. C, Subcellular localization of OsGZ7 protein in rice protoplasts. The constructs of green fluorescent protein (GFP) and OsGZ7-GFP were transiently expressed in the protoplasts prepared from rice. GFP was visualized using a laser scanning microscope (LSM 980, Zeiss). Scale bars, 10 μm. D-J, Phenotype (D), shoot length (E), shoot dry weight (F), root dry weight (G), Zn content in shoot (H), Zn content in root (I), and Zn concentration in brown rice (J) of wild type (WT), nockout (KO-1 and KO-2), and overexpression (OE1 and OE2) lines of OsGZ7 under Zn-normal (0.4 μmol/L) and Zn-deficient (0.04 μmol/L) conditions after 17 d of treatment. K, Number of differentially expressed genes (DEGs) between WT and KO lines under Zn-deficient conditions. L, Venn diagram showing the number of Zn-normal and -deficiency-responsive DEGs between WT and KO lines under Zn-normal and Zn-deficient conditions. M, Kyoto Encyclopedia of Gene and Genomes (KEGG) analysis of upregulated genes under Zn-deficient conditions. In E-J, data are mean ± SD (n ≥ 3). * and ** above bars indicate significant differences between WT and each of the KO and OE lines at the 0.05 and 0.01 levels by Tukey’s HSD test.