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通过PCR方法从水稻“矮子占”和“南特号”的基因组DNA中分离出赤霉素20-氧化酶基因rga5,其编码区含1119 bp,其编码蛋白与已报道的水稻赤霉素20-氧化酶基因OsGA20ox相比有11个氨基酸不同,其中9个氨基酸为移码差异,通过基因枪法将该基因的正、反义表达质粒pSrga5和pArga5转化水稻“中花8号”,获得了生物学性状有明显变异的转基因植株,正义转化表现出植株增高、叶片和穗变长、穗粒数增加等特征,但花期未受明显的影响;而反义植株表现出明显的“矮化”和“早花”,且出现植株细弱、叶色加深、叶片和穗变短等特征,转基因水稻的分析结果表明,rga5正、反义外源基因已分别整合到水稻基因组中,并有效表达;正义植株体内的GA1平均增加约50％,反义则降低至对照组的约10％.结果表明,rga5是水稻赤霉素合成代谢途径中起关键作用的赤霉素20-氧化酶基因,该序列中11个氨基酸的变异并不影响其功能,正义表达可增强体内GA的合成,促使植株生长,增加株高;反义转化则抑制了内源GA的合成及植株生长,导致植株明显的矮化.
The GA3 gene of GA3 was isolated from the genomic DNA of “Dwarf Zhanzi” and “Nantes” by PCR and the coding region contained 1119 bp. The coding region of GA35 was related to the reported gibberellin 20 - oxidase gene OsGA20ox compared to 11 amino acids, of which 9 amino acids for the frame shift difference, the gene by positive and negative sense of the gene expression plasmid pSrga5 and pArga5 rice “Zhonghua 8” to obtain a biological Transgenic plants with significant variation in their traits exhibited the characteristics of increased plant height, longer leaves and spikes and increased grains per spike, but the florescence was not significantly affected; while the antisense plants showed obvious “dwarfing” and “Early flowering”, and the characteristics of thin plants, deepened leaf color and shorter leaves and ears appeared. The analysis of transgenic rice showed that positive and antisense rga5 genes had been integrated into rice genome and expressed effectively. The average GA1 increased about 50% in plant and the antisense decreased to about 10% in control group.The results showed that rga5 was the gibberellin 20-oxidase gene that plays a key role in the gibberellin metabolic pathway, In 11 ammonia Acid mutation does not affect its function, it can enhance the sense expression of GA synthesis in vivo, promote plant growth, increased plant height; antisense suppressed the endogenous conversion of GA synthesis and plant growth, resulting in dwarfed plants significantly.