1个新水稻组成型启动子的克隆与功能鉴定

根据基因芯片数据库和RT-PCR验证得到1个高活性的水稻组成型表达基因(TIGR Locus:LOC-Os07g34589),用PCR技术从籼稻品种明恢63基因组中克隆得到其上游启动子PSUI1,长度为1 941bp;将其与β-glucuronidase(GUS)报告基因融合构建植物表达载体DX2181b-PSUI1,利用玉米Ubiquitin启动子融合GUS报告基因构建表达载体DX2181b-PUbi作为对照,通过根癌农杆菌(Agrobacterium tumefaciens)介导法将DX2181b-PSUI1和DX2181b-PUbi转化粳稻品种中花11。组织化学染色表明,含DX2181b-PSUI1的转基因植株中,GUS基因在幼苗期叶片、叶鞘、根,抽穗期叶片、叶鞘、茎秆、颖壳、雄蕊和成熟期的叶片、叶鞘、茎秆、胚、胚乳中均有表达,说明PSUI1为组成型启动子。对GUS表达活性进行定量分析表明,PSUI1启动子的活性约为玉米Ubiquitin启动子活性的1/3～1/2,但是PSUI1表现出了更好的表达稳定性。

Isolation and characterization of a novel rice constitutive promoter

A constitutive gene(TIGR Locus:Loc_Os07g34589) was obtained by combining our lab’s database with RT-PCR verification.A novel rice constitutive promoter named PSUI1 with 1 941 bp was isolated using the genomic DNA of the indica rice variety Minghui63 based on a PCR method.To determine the expression strength and pattern of the novel constitutive promoter,the cloned promoter was fused with the β-glucuronidase(GUS) reporter gene to form plant expression vector DX2181b-PSUI1,and the maize ubiquitin promoter was also fused with GUS reporter gene to form expression vector DX2181b-PUbi as the control.Both DX2181b-PSUI1 and DX2181b-PUbi were transformed into japonica rice variety Zhonghua11 by Agrobaeterium-mediated method.GUS staining showed that GUS gene was expessed in different rice tissues and organs at different growth stages of DX2181b-PSUI1 transgenic rice plants.Quantitative analysis of GUS activity showed that the strength of maize ubiquitin promoter was 2-3 times higher than that of the PSUI1 promoter,but the expression of PSUI1 promoter exhibited much more stable than that of maize Ubiquitin promoter between transgenic rice lines.