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. 2019 Aug 3;69(3):429–438. doi: 10.1270/jsbbs.19014

Fig. 3.

Fig. 3

Molecular confirmation of transgenic rice lines expressing TrGDH-FLAG. (A) Schematics of plant expression vector pCAMBIA1301-Ubi::TrGDH-FLAG. HPT hygromycin resistance gene, Tnos terminator of nopaline synthase gene (nos), FLAG tag sequence, AsRed red fluorescence protein gene, 35S CaMV 35S promoter, Ubi ubiquitin promoter derived from Zea mays, LB left border, RB right border, and TrGDH NADP(H)-GDH gene derived from Trichurus. (B) Relative expression level analysis of TrGDH in the control and TrGDH transgenic rice lines by semi-quantitative RT-PCR. Two independent transgenic rice lines, Ubi::TrGDH-4 and Ubi::TrGDH-13, were used for analysis. OsActin1 was amplified as a control for mRNA levels. (C) Western blotting analysis of TrGDH-FLAG in the control and TrGDH transgenic rice lines using anti-FLAG antibody (1:5000). Poceacu staining was used as a loading control. (D) Real-time quantitative PCR analysis of OsGDH1, OsGDH2, OsGDH3, OsGDH4 in the control and TrGDH transgenic rice lines. OsActin1 was amplified as a control for mRNA levels. Triplicate quantitative assays were performed on each cDNA sample. * indicates significant differences at p < 0.05. ** indicates significant differences at p < 0.01.