School of Biotechnology and Environmental Engineering, Myongji University, Yongin 449-728, Korea.
Planta. 2012 Jun;235(6):1397-408. doi: 10.1007/s00425-011-1582-x. Epub 2012 Jan 3.
We have previously characterized the constitutively active promoters of the APX, PGD1 and R1G1B genes in rice (Park et al. 2010 in J Exp Bot 61:2459-2467). To have potential crop biotechnology applications, gene promoters must be stably active over many generations. In our current study, we report our further detailed analysis of the APX, PGD1 and R1G1B gene promoters in various organs and tissues of transgenic rice plants for three (T₃₋₅) homozygous generations. The copy numbers in 37 transgenic lines that harbor promoter:gfp constructs were determined and promoter activities were measured by real-time qPCR. Analysis of the 37 lines revealed that 15 contained a single copy of one of the three promoter:gfp chimeric constructs. The promoter activity levels were generally higher in multi-copy lines, whereas variations in these levels over the T₃₋₅ generations studied were observed to be smaller in single-copy than in multi-copy lines. The three promoters were further found to be highly active in the whole plant body at both the vegetative and reproductive stages of plant growth, with the exception of the APX in the ovary and R1G1B in the pistil and filaments where zero or very low levels of activity were detected. Of note, the spatial activities of the PGD1 promoter were found to be strikingly similar to those of the ZmUbi1, a widely used constitutive promoter. Our comparison of promoter activities between T₃, T₄ and T₅ plants revealed that the APX, PGD1 and R1G1B promoters maintained their activities at comparable levels in leaves and roots over three homozygous generations and are therefore potentially viable alternative promoters for crop biotechnology applications.
我们之前已经对水稻中 APX、PGD1 和 R1G1B 基因的组成型活性启动子进行了特征描述（Park 等人，2010 年在《实验植物学杂志》上，2459-2467）。为了具有潜在的作物生物技术应用，基因启动子必须在许多代中保持稳定的活性。在我们目前的研究中，我们报告了对 APX、PGD1 和 R1G1B 基因启动子在转基因水稻植株的各种器官和组织中的进一步详细分析，这些植株为三个（T₃₋₅）纯合世代。对携带启动子：gfp 构建体的 37 个转基因系的拷贝数进行了测定，并通过实时 qPCR 测量了启动子活性。对 37 个系的分析表明，有 15 个系含有三种启动子：gfp 嵌合构建体之一的单个拷贝。多拷贝系中的启动子活性水平通常较高，而在研究的 T₃₋₅ 世代中，这些水平的变化在单拷贝系中比在多拷贝系中要小。还发现，这三个启动子在整个植物体内的营养和生殖阶段都具有高度的活性，除了在卵巢中的 APX 和在雌蕊和花丝中的 R1G1B 之外，在这些部位检测到的活性为零或非常低。值得注意的是，PGD1 启动子的空间活性与广泛使用的组成型启动子 ZmUbi1 非常相似。我们对 T₃、T₄ 和 T₅ 植株之间启动子活性的比较表明，APX、PGD1 和 R1G1B 启动子在三个纯合世代的叶片和根部保持相似的活性水平，因此是作物生物技术应用的潜在可行替代启动子。
