玉米盐诱导型启动子克隆及其植物表达载体构建 Isolation of Salt-Inducible Promoter in Maize and Construction of Plant Expression Vector

ISSN： 2168-5673

Source： Botanical Research, Vol.01, Iss.02, 2012-07, pp. : 30-35

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Abstract

首先提取B73玉米(Zea mays)的基因组DNA，以此为模板利用PCR的方法克隆Zea mays salt-inducible protein kinase(STY2)基因上游1675 bp，将其命名为STY。从网站PlantCARE上的启动子预测工具在线对克隆到的启动子进行分析，结果表明：STY启动子中含有多种调控元件，典型的调控元件有：TATA-box，CAAT-box等等。另外还有2个盐诱导顺式作用元件GT1-motif和一些冷、干旱胁迫诱导有关的顺式作用元件，以及功能未知的元件。然后将克隆到的基因与植物双元表达载体pCAMBIA1301连接，并将其转入根癌农杆菌EHA105中，然后利用农杆菌介导法转化烟草，经过GUS组织化学染色验证STY的诱导功能。研究表明：STY启动子片段具有启动活性，为解决东北地区农业生产中严重的多元逆境胁迫问题开辟了有效途径。 The upstream nucleotide sequence of maize salt-inducible protein kinase, named STY, was isolated from the genomic DNA of maize by PCR. Promoter sequence analysis by PlantCARE showed that it had some typical cis-elements, including TATA box and CAAT box. In addition, there were two salt responsive elements GT1-motif, various phytohormone responsive elements and stress responsive elements in the promoter sequence. The fusion gene of STY promoter and GUS were constructed and named pCAM-STY. The vector was trans-formed into Agrobacterium tumefaciens. The vector containing STY promoter was transferred into tobacco by Agrobacterium tumefaciens mediated method, function of STY promoter was analyzed by identifying activation of GUS using histochemistry staining. The results indicated that the promoter activated expression of GUS re-port gene, It laid the foundation to resolve the serious multi-stress problems of agriculture in northeast of China.