发根农杆菌介导丹参牻牛儿基牻牛儿基焦磷酸合酶1基因RNA干扰载体的转化

目的：利用发根农杆菌ACCC10060介导丹参牻牛儿基牻牛儿基焦磷酸合酶1基因（SmGGPS1）RNA干扰（RNAi）载体转化丹参叶片,生成SmGGPS1的RNAi转基因毛状根。方法：根据已克隆到的SmGGPS1特异区域设计并合成2段RNAi序列,分别插入RNAi双元载体pK7GWIWG2D（Ⅱ）中,构建2个含卡那霉素（Kan）和绿色荧光蛋白（GFP）双筛选标记的植物表达载体pK7GWIWG2D（Ⅱ）-SmGGPS1-RNAi2和pK7GWIWG2D（Ⅱ）-SmGGPS1-RNAi3;利用带有上述2个RNAi载体的发根农杆菌ACCC10060侵染丹参叶片,诱导生成转基因毛状根;通过Kan抗性筛选和GFP绿色荧光观察统计转化率。结果：分别得到SmGGPS1-RNAi2和SmGGPS1-RNAi3转基因毛状根301根和399根,平均转化率为60.34%。结论：首次建立了发根农杆菌介导的外源基因转化丹参的体系。

Agrobacterium rhizogenes-Mediated Transformation of Salvia miltiorrhiza Bunge Against the RNAi Vectors of Geranylgeranyl Pyrophosphate Synthase 1 Gene

Objective： Mediated by Agrobacterium rhizogenes ACCC10060, the RNA interference（RNAi） vectors against Salvia miltiorrhiza Bunge geranylgeranyl pyrophosphate synthase 1（SmGGPS1） gene were transformed into Danshen＇s leaves, and the RNAi transgenic hairy roots were generated. Methods： According to the obtained SmGGPS1 specific regions, the two RNAi sequences were designed and inserted into the binary RNAi vector pK7GWIWG2D（ Ⅱ ）, then pKTGWIWG2D（Ⅱ）- SmGGPS1-RNAi2 and pKTGWIWG2D（Ⅱ）-SmGGPS1-RNAi3, the two plant expression vectors with Kan and GFP markers were constructed. Danshen＇s leaves were infected with A.rhizogenes ACCC10060 harboring the two RNAi vectors, and then the hairy roots were induced. The infected hairy roots were screened by Kan resistance, and the green fluorescent of GFP in the cut hairy roots was observed further to calculate the transformation rate. Results： The number of SmGGPS1-RNAi2 and SmGGPS1-RNAi transgenic hairy roots were 301 and 399 respectively, with the average transformation rate of 60.34%. Conclusion： The A.rhizogenes-mediated foreign gene transformation system of Danshen was firstly established.