水稻基因启动子OsBTF3p的克隆和启动活性分析

 【目的】分子克隆水稻基因OsBTF3启动子片段，明确其对靶基因表达的启动作用，为抗病转基因水稻研究提供理论依据和启动子元件材料。【方法】对OsBTF3编码区上游1387 bp的启动子（OsBTF3p）序列进行了克隆和序列分析，构建了OsBTF3p∷GUS融合基因植物表达载体pCAM-OsBTF3p，利用农杆菌介导的水稻遗传转化，获得了39株OsBTF3p∷GUS转基因植株，对OsBTF3p进行了启动活性、组织特异性及病原菌诱导性分析。【结果】分子克隆了OsBTF3p片段，其序列与GenBank中的已知序列一致。在转基因水稻愈伤组织中能够检测到GUS活性，表明该启动子具有启动活性。在转基因水稻叶片维管束组织和根部组织能检测到GUS活性。水稻白叶枯病菌(Xoo)侵染后OsBTF3p驱动的GUS活性明显地上调表达。【结论】OsBTF3p具有驱动GUS基因表达的启动活性、组织表达特异性和病原菌诱导性。

Molecular Cloning and Activity Analysis of a Rice Gene Promoter OsBTF3p

【Objective】 To make a cloning and functional analysis of OsBTF3p, a rice gene promoter directing GUS expression. The functional analysis of OsBTF3p is helpful to understand the mechanisms of gene expression and to utilize in generation of transgenic plants. 【Method】 A 1378bp promoter of an inducible rice gene OsBTF3 upon infection by Xanthomoas oryzae pv. oryzae (Xoo) was amplified by PCR and named as OsBTF3p. OsBTF3p fused in frame with the gus reporter gene and the resulting construct (OsBTF3p::GUS) was transformed into the rice calli through Agrobacterium-mediated transformation. Thirty-nine transgenic plants of rice （T0 generation） were successfully obtained. The promoter activity, tissue specificity and pathogen inductivity were analyzed as well. 【Result】 The cloned DNA sequence of OsBTF3p is same as that present in the GenBank database. GUS activity can be detected in the OsBTF3p::GUS -transformed but non-transgenic calli (control). GUS expression was also observed in the OsBTF3p::GUS-transformed vascular tissues of leaves and roots. The transgene OsBTF3p::GUS in rice is up-regulated upon Xoo infection. 【Conclusion】 OsBTF3p might function effectively in promoting GUS expression in transgenic rice plants in tissue-specific and pathogen-inducible fashions.