Qu Le Qing, Takaiwa Fumio
Department of Plant Biotechnology, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba, Ibaraki 305-8602, Japan.
Plant Biotechnol J. 2004 Mar;2(2):113-25. doi: 10.1111/j.1467-7652.2004.00055.x.
Using stable transgenic rice plants, the promoters of 15 genes expressed in rice seed were analysed for their spatial and temporal expression pattern and their potential to promote the expression of recombinant proteins in seeds. The 15 genes included 10 seed storage protein genes and five genes for enzymes involved in carbohydrate and nitrogen metabolism. The promoters for the glutelins and the 13 kDa and 16 kDa prolamins directed endosperm-specific expression, especially in the outer portion (peripheral region) of the endosperm, whilst the embryo globulin and 18 kDa oleosin promoters directed expression in the embryo and aleurone layer. Fusion of the GUS gene to the 26 kDa globulin promoter resulted in expression in the inner starchy endosperm tissue. It should be noted that the 10 kDa prolamin gene was the only one tested that required both the 5' and 3' flanking regions for intrinsic endosperm-specific expression. The promoters from the pyruvate orthophosphate dikinase (PPDK) and ADP-glucose pyrophosphorylase (AGPase) small subunit genes were active not only in the seed, but also in the phloem of vegetative tissues. Within the seed, the expression from these two promoters differed in that the PPDK gene was only expressed in the endosperm, whereas the AGPase small subunit gene was expressed throughout the seed. The GUS reporter gene fused to the alanine aminotransferase (AlaAT) promoter was expressed in the inner portion of the starchy endosperm, whilst the starch branching enzyme (SBE1) and the glutamate synthase (GOGAT) genes were mainly expressed in the scutellum (between the endosperm and embryo). When promoter activities were examined during seed maturation, the glutelin GluB-4, 26 kDa globulin and 10 kDa and 16 kDa prolamin promoters exhibited much higher activities than the others. The seed promoters analysed here exhibited a wide variety of activities and expression patterns, thus providing many choices suitable for various applications in plant biotechnology.
利用稳定的转基因水稻植株，对在水稻种子中表达的15个基因的启动子进行了分析，以研究其空间和时间表达模式以及在种子中促进重组蛋白表达的潜力。这15个基因包括10个种子贮藏蛋白基因和5个参与碳水化合物和氮代谢的酶基因。谷蛋白以及13 kDa和16 kDa醇溶蛋白的启动子指导胚乳特异性表达，特别是在胚乳的外部（周边区域），而胚球蛋白和18 kDa油质蛋白启动子指导在胚和糊粉层中的表达。GUS基因与26 kDa球蛋白启动子融合导致在内胚乳淀粉组织中表达。应当指出，10 kDa醇溶蛋白基因是唯一测试的需要5'和3'侧翼区域才能实现内在胚乳特异性表达的基因。丙酮酸磷酸双激酶（PPDK）和ADP - 葡萄糖焦磷酸化酶（AGPase）小亚基基因的启动子不仅在种子中具有活性，而且在营养组织的韧皮部中也具有活性。在种子内部，这两个启动子的表达有所不同，因为PPDK基因仅在胚乳中表达，而AGPase小亚基基因在整个种子中表达。与丙氨酸转氨酶（AlaAT）启动子融合的GUS报告基因在内胚乳淀粉的内部表达，而淀粉分支酶（SBE1）和谷氨酸合酶（GOGAT）基因主要在盾片（胚乳和胚之间）中表达。当在种子成熟过程中检查启动子活性时，谷蛋白GluB - 4、26 kDa球蛋白以及10 kDa和16 kDa醇溶蛋白启动子表现出比其他启动子高得多的活性。这里分析的种子启动子表现出广泛的活性和表达模式，从而为植物生物技术中的各种应用提供了许多合适的选择。
