| 摘要: |
| 【目的】 构建具有大豆蛋白酶抑制剂BBi基因反向重复结构的ihpRNA种子特异性表达载体，并将其转入根癌农杆菌中，为大豆品质改良奠定基础。【方法】 采用PCR技术克隆大豆蛋白酶抑制剂BBi基因的正义和反义片段、大豆种子特异性启动子7αP和作为内含子的GFP基因片段，分别连入克隆载体pMD18-T Vector中。然后根据植物中ihpRNA原理，以植物表达载体pCAMBIA1301为基础，将组成RNAi 载体的4个目的片段分别连入其中，然后利用冻融法转化根癌农杆菌，并进行PCR鉴定。【结果】 PCR扩增得到组成RNAi 载体的4个目的片段，分别构建成重组克隆载体和 ihpRNA种子特异性表达载体p7αP-GFP-BBiS，并转化得到含有p7αP-GFP-BBiS的农杆菌EHA101和EHA105。【结论】 成功构建了具有BBi基因反向重复结构的ihpRNA种子特异性表达载体p7αP-GFP-BBiS，BBi基因的ihpRNA表达框架成功转入到农杆菌中。 |
| 关键词: 大豆 蛋白酶抑制剂BBi基因 ihpRNA表达载体 种子特异性启动子 |
| DOI： |
| 分类号: |
| 基金项目:教育部博士点基金项目（20070193005） |
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| Construction of ihpRNA seed-specific expressed vector of Bowman-Birk inhibitor gene from soybean |
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FU Yong-ping,ZHANG Jun,WANG Pi-wu,ZHOU Hai-tao and Qu Jing,FU Yong-ping,ZHANG Jun,WANG Pi-wu,ZHOU Hai-tao and Qu Jing,FU Yong-ping,ZHANG Jun,WANG Pi-wu,ZHOU Hai-tao and Qu Jing,FU Yong-ping,ZHANG Jun,WANG Pi-wu,ZHOU Hai-tao and Qu Jing,FU Yong-ping,ZHANG Jun,WANG Pi-wu,ZHOU Hai-tao and Qu Jing
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| Abstract: |
| 【Objective】 The ihpRNA seed-specific expressed vector of Bowman-Birk inhibitor gene from soybean was constructed,and then introduced into Agrobacterium tumefaciens to lay a foundation for quality improvement of soybean.【Method】 In this paper,the BBi gene,soybean seed-specific promotor 7αP and the fragment of GFP gene which were used as intron were isolated by PCR method,and then inserted into pMD18-T Vector respectively.The ihpRNA seed-specific expressed vector p7αP-GFP-BBiS with inverted repeats of BBi genes,seed-specific promotor 7αP and GFP gene was constructed based on the vector of pCAMBIA1301.p7αP-GFP-BBiS was introduced into EHA101 and EHA105 of A.tumefaciens respectively.【Result】 The BBi gene,soybean seed-specific promotor 7αP and the fragment of GFP gene were cloned,and the four recombinant cloned vectors and the ihpRNA seed-specific expressed vector p7αP-GFP-BBiS were constructed.The EHA101 and EHA105 which had p7αP-GFP-BBiS were obtained.【Conclusion】 The ihpRNA seed-specific expressed vector p7αP-GFP-BBiS was constructed with inverted repeats of BBi gene,seed-specific promotor 7αP and GFP gene,and the RNAi constructs were transformed into A.tumefaciens. |
| Key words: soybean Bowman-Birk inhibitor gene ihpRNA expression vector seed-specific promotor |
