双价抗虫转基因水稻的育成及初步鉴定  

Breeding and preliminary identification of binary insectresistant transgenic rice

作　　者：崔彦芹[1] 李尚伟[2] 张丽萍[2] 罗洪发[1] 李云峰[3] 查仁明[1] 

机构地区：[1]贵州大学农学院,贵阳550025 [2]贵州大学昆虫研究所/贵州山地农业病虫害重点实验室,贵阳550025 [3]西南大学水稻研究所/转基因植物与安全控制重庆市重点实验室,重庆400716

出　　处：《南方农业学报》2016年第2期169-173,共5页Journal of Southern Agriculture

基　　金：贵州省科技计划项目(黔科合NY字[2011]3007号)

摘　　要：【目的】构建双价抗虫基因表达载体并转化水稻,为选育广谱、高抗转基因抗虫水稻提供技术参考。【方法】构建半夏凝集素基因(pta)和苏云金杆芽孢菌基因(Bt)两个抗虫基因的双价表达载体p CAMBIA1301-Bt-pta,利用大肠杆菌DH5α和根癌农杆菌LBA4404转化水稻恢复系辐恢838,同时对转化植株进行GUS染色及PCR鉴定。【结果】以构建的双价植物表达载体p CAMBIA1301-Bt-pta为模板,PCR扩增到pta基因的部分片段,大小为807 bp,且该载体经HindⅢ酶切后,能得到15854 bp的中间载体p CAMBIA1301-Bt片段和2046 bp的pta基因片段。对转化植株进行GUS染色,得到3株转基因阳性植株,经PCR鉴定结果显示外源基因已成功导入水稻。【结论】通过构建双价抗虫基因表达载体并转化水稻,成功获得的转基因植株可用于水稻抗虫转基因恢复系及组合的研究,以提高其抗病虫害能力。【Objective】The present experiment was conducted to construct a bivalent plant expression vector carrying two insect-resistant genes and transfer it into rice, in order to lay foundation for obtaining broad-spectrum and high insect-resistant transgenic rice. 【Method】A bivalent plant expression vector（p CAMBIA1301-Bt-pta） carrying two insect-resistant genes viz., Pinellia ternate agglutinin gene（pta） and Bacillus thuringiensis gene（Bt） was constructed, and then transferred into rice restorer Fuhui 838. Then, the transferred plants were identified by GUS staining and PCR detection.【Result】The results showed that part pta gene fragment with 807 bp in length was amplified using constructed bivalent plant expression vector（p CAMBIA1301-Bt-pta） as template. And after p CAMBIA1301-Bt-pta vector were digested by HindⅢ, based on electrophoresis analysis, there were a big fragment（15854 bp in length） and a small fragment（2046 bp in length） to be found, which were the main fragment of intermediate vector p CAMBIA1301-Bt and pta gene, respectively.These results showed that pta gene had been connected to intermediate vector p CAMBIA1301-Bt. Furthermore, the results of GUS staining assay showed that only 3 plants was positive, and the PCR amplification results showed that the exogenous gene had been transferred into rice. 【Conclusion】By constructing bivalent expression vector and transfering insect-resistant genes into rice, the transgenic plants were obtained successfully, which can be used to study insect-resistant transgenic restoration lines and combination, so as to increase its resistance to insects and diseases.

关 键 词：BT基因 pta基因 载体构建 转基因水稻 鉴定 

分 类 号：S511.035.3[农业科学—作物学]