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当前位置: 首页 > 文章 > pCB-zeolin-GFP表达载体的构建及瞬时表达 中国农学通报 2012,28 (3) 233-239
Position: Home > Articles > Construction of Plant Expression Vector pCAMBIAl302-zeolin-GFP and the Transient Expression Chinese Agricultural Science Bulletin 2012,28 (3) 233-239

pCB-zeolin-GFP表达载体的构建及瞬时表达

作  者：
张玉;白史且;李聪;李达旭;邓永昌
单  位：
中国农业科学院北京畜牧兽医研究所;四川省草原科学研究院
关键词：
绿色荧光蛋白;zeolin基因;载体构建;瞬时表达
摘  要：
为利用荧光蛋白基因GFP检测外源基因在转基因植株中的表达和定位,构建含有GFP基因的植物表达载体pCB-zeolin-GFP。在目的基因的开放阅读框(ORF)两端设计引物,并引入酶切位点和保护碱基,用PCR方法从pDHA扩增得到zeolin基因的全长,克隆到中间载体pMD18-T,分别用NcoⅠ和BglⅡ2种限制性内切酶酶切重组质粒和经过改良的pCAMBIAI1302植物表达载体,经回收、连接、转化、鉴定后,利用基因枪转化法将重组载体转入洋葱表皮细胞,通过共聚焦显微镜检测绿色荧光蛋白在洋葱表皮细胞中的瞬时表达。构建了zeolin基因与绿色荧光蛋白(GFP)融合的植物表达载体pCB-zeolin-GFP,并在洋葱中得到了表达。构建的融合植物表达载体pCB-zeolin-GFP正确,该载体的成功构建为今后进行基因转移、基因功能研究及培育新品种奠定了基础。
译  名：
Construction of Plant Expression Vector pCAMBIAl302-zeolin-GFP and the Transient Expression
作  者：
Zhang Yu 1 , Bai Shiqie 1 , Li Cong 2 , Li Daxu 1 , Deng Yongchang 1 (1 Sichuan Grassland Science Academy , Xipu Chengdu 611731; 2 Institute of Animal Science, Chinese Academy of Agricultural Science , Beijing 100094)
关键词：
green fluorescent protein (GFP); zeolin gene; vector construction; transient expression
摘  要：
To utilize fluorescin gene GFP to detect the express and localization of exogenous gene, plant expression vector pCAMBIAl302-zeolin with a green fluorescent protein gene would be constructed. The total length sequence of zeolin gene in pDHA plasmid was amplified by PCR. The fragment was cloned into pMD18-T middle vector, and a new recombined vector named pMD18-T-zeolin was obtained. A new plant expression vector named pCB-GFP-zeolin was constructed after cutting two vector pMD18-zeolin and pCAMBIAI1302 with restriction enzymes, subsequently reclamation, ligation, transformation and identification. Then the recombined plant expression vector pCAMBIAl302-zeolin with a green fluorescent protein gene was transformed into epidemic cells of onion by gene gun method and was detected by confocal microscopy. The recombined plant expression vector pCAMBIAl302-zeolin with a green fluorescent protein gene had been constructed and the fusion gene was transient expressed. It was suggested that the recombined plant expression vector pCAMBIAl302-zeolin with a green fluorescent protein gene was successful. Construction of the plant expression vector might play an important role in genetic transformation, the gene functional identification and breeding of new varieties.

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