拟南芥PhrIP1基因超表达、GFP和RNAi载体的构建  

Construction of Plant Over-expression,GFP and RNAi Vectors of PhrIP1 of Arabidopsis Thaliana

作　　者：马立安[1] 张忠明[2] 

机构地区：[1]长江大学生命科学学院,湖北荆州434025 [2]华中农业大学农业微生物学国家重点实验室,湖北武汉430070

出　　处：《长江大学学报（自科版）（中旬）》2007年第3期73-76,99,共5页Journal of Yangtze University(Nature Science Edition)

基　　金：国家自然科学基金项目(30070370);美国国家科学基金项目(NSF-MCB0548525)

摘　　要：根据编码拟南芥成膜素相关蛋白(PhrIP1)基因cDNA全序列设计超表达引物,以1～1000bp之间序列设计GFP引物和序列内部第24～240bp之间序列设计RNAi引物,以pMD18-T-PhrIP1为模板,用PCR方法分别扩增出1.8kb、1.0kb和0.216kb的片段,分别克隆至双元表达载体、GFP载体和RNAi载体上,得到了植物超表达载体pBI-PhrIP1、GFP载体pMON-GFP-PhrIP1和RNAi载体Hellsgate2-PhrIP1。用电转化法,将这些重组质粒导入农杆菌GV3101菌株中,PCR扩增结果表明所构建的植物超表达载体pBI-PhrIP1、GFP融合载体pMON-GFP-PhrIP1和RNAi载体Hellsgate2-PhrIP1已导入农杆菌。A 1.8 kb,a 1.0 kb and a 0.216 kb fragments were amplified respectively from complete,nucleotides 1 to 1 000 and nucleotides 24 to 240 of the cDNA sequence of PhrIP1 gene that encodes a PhrIP1 interacting with phragmoplastin.The produced fragments were respectively introduced to the binary expressed vector pBI121 to produce pBI-PhrIP1 and to the GFP expressed vector pMON18342 to produce pMON-GFP-PhrIP1 and to the RNAi vector Hellsgate2 to produce Hellsgate2-PhrIP1,and then these plasmids were respectively mobilized into Agrobacterium tumefaciens strain GV3101 by electrotransformation.The results of PCR amplifications showed that the PhrIP1 gene fragment,GFP gene fragment and RNAi gene fragment of PhrIP1 were introduced into pBI121,pMON18342 and Hellsgate2,and the generated pBI-PhrIP1,pMON-GFP-PhrIP1 and Hellsgate2-PhrIP1 were respectively transferred into the Agrobacterium tumefaciens successfully.

关 键 词：拟南芥PhrIP1基因 超表达载体 RNAI载体 GFP融合载体 电转化法 

分 类 号：Q784[生物学—分子生物学]