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水稻OsTLP12基因启动子GUS表达载体的构建与转化
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Abstract:
植物中类甜蛋白TLPs不仅参与宿主抵御真菌入侵的过程，还参与植物对非生物胁迫的防御。本试验采用PCR技术从籼稻特青中克隆了OsTLP12基因上游2000 bp的启动子片段，并分析了其中所含的顺式作用元件。随后，构建了与GUS报告基因融合的表达载体，并通过农杆菌介导的转化方法将融合表达载体导入水稻中，随后进行了转化植株的筛选，并对转基因植株进行了GUS染色分析。本研究得到以下结果：1) 成功克隆了水稻中OsTLP12基因的启动子片段，并将其与含有GUS报告基因的表达载体融合，获得了融合表达载体；2) OsTLP12基因启动子区域含有多个生物和非生物逆境反应相关的顺式元件；3) 成功将融合表达载体转化进入水稻中，并筛选出阳性转基因幼苗；4) 对转基因幼苗进行了GUS染色，以检测OsTLP12基因启动子驱动的GUS表达情况。
Plant thaumatin-like proteins (TLPs) are involved not only in host defense against fungal invasion but also in defense against abiotic stresses. In this experiment, the upstream 2000 bp promoter fragment of the OsTLP12 gene was cloned from Oryza sativa ‘Nipponbare’ using PCR technology, and the cis-acting elements contained therein were analyzed. Subsequently, a fusion expression vector containing the GUS reporter gene was constructed, and the fusion expression vector was introduced into rice via Agrobacterium-mediated transformation. The transformed plants were then screened, and GUS staining analysis was performed on the transgenic plants. The following results were obtained: 1) The promoter fragment of the OsTLP12 gene in rice was successfully cloned, and it was fused with the expression vector containing the GUS reporter gene to obtain the fusion expression vector; 2) The promoter region of the OsTLP12 gene contains multiple cis-acting elements related to biotic and abiotic stress responses; 3) The fusion expression vector was successfully transformed into rice, and positive transgenic seedlings were screened out; 4) GUS staining was performed on transgenic seedlings to detect the expression driven by the OsTLP12 gene promoter.
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