紫花苜蓿MsFPS基因超表达载体的构建及烟草转化

Construction of Super-expression Plasmid of MsFPS Gene from Medicago sativa L. and Transformation of Tobacco

采用RT-PCR技术获得紫花苜蓿的法呢基焦磷酸合成酶基因,并连接到含有35S启动子和GUS基因的载体pBI121上,成功构建植物表达载体pBI-FPS。用根癌农杆菌介导法将其转入烟草中,共获得7株卡那霉素抗性苗,对其中的4株进行PCR检测,证明目的基因已经整合到烟草基因组中。进一步对这4株进行RT-PCR和组织化学染色法检测,初步证实目的基因在烟草中可以表达,说明已成功获得能够表达MsFPS基因的转基因烟草。

By RT-PCR technology, the cDNA of FPS（Farnesyl phosphate synthase） gene ot altalla lvleazcago sativa L. ） was obtained, and it was inserted into plasmid pBI121 that contains promoter 35S and GUS gene. A plant super expressed vector of MsFPS gene from alfalfa was constructed successfully and was transferred to tobacco by the Agrobacteriurn mediated transformation system. Seven resistant plants to kanamycin were obtained. PCR analysis of four of them showed that the target gene had been integrated into the genomes of the tobacco. And expression of the MsZFN gene had been Confirmed in four tobaco plants by RT-PCR and GUS gene expression. As a result, transgenic tobacco in which MsFPS could be expressed had successfully been obtained.