蜡梅水通道蛋白基因CpAQP1转录水平变化及其启动子克隆

Changes in Transcription Level of the Aquaporin Gene CpAQP1 from Chimonanthus praecox and the Cloning of Its Promoter

利用荧光定量PCR技术对蜡梅花发育的不同组织、不同花期和不同胁迫下的水通道蛋白基因CpAQP1的表达量进行分析.结果表明:CpAQP1在盛开期中瓣的表达量明显高于其他组织器官,在高温、高盐、ABA胁迫下表达量变化明显,说明该基因具有组织表达差异性及能够对环境胁迫做出响应.同时利用hi-TAIL PCR方法克隆获得蜡梅水通道蛋白基因CpAQP1上游启动子1082bp,命名为CpAQP1pro(GenBank登录号为:JQ952563).该序列具有典型的基本元件TATA-box、CAAT-box及与胁迫相关的元件和光应答元件.将克隆的启动子CpAQP1pro替换pBI121中的CaMV35s启动子,构建植物表达载体pBI121-CpAQP1pro,通过农杆菌转化烟草,稳定表达结果说明该启动子具有驱动GUS报告基因表达的活性.

In order to explore the relationship between CpAQP1 expression level and stress in plants, the RT-PCR method was used to analyze the expression of CpAQP1 in different organs of Chirnonanthus praecox at different developmental stages and under different stress conditions in this research. The results showed that CpAQP1 was expressed at a higher level in the petal than in other organs during the full blooming stage of the plant, and it showed no significant changes under abiotic stresses （high temperature, salinity and ABA）, indicating that CpAQP1 may have the organizational differences in its expression and make responses to abiotic stresses. Meanwhile, a promoter fragment of 1082 bp upstream from the 5＇ upper of CpAQP1 was cloned from the genomic DNA of C. praecox by hiTAIL-PCR. This sequence was thereafter designated as CpAQPlpro （GenBank accession No. JQ952563）. Bioinformatics analysis revealed that the sequence contained the basic cis elements, such as TATA-box and CAAT box, and some elements involved in the plant abiotic stress and light responses. The promoter CaMV35s of pBI121 was replaced by the cloned promoter CpAQPlpro, and then the plant expression vector pBI121-CpAQPlpro was successfully established. The result of stable expression by the Agrobacterium tumefaciens-mediated method indi cated that the promoter fragment had the function to drive the expression of the GUS （b-glucuronidase） reporter gene.