鄂马铃薯3号遗传转化体系的构建与表达

Construction and expression of genetic transformation system of Hubei potato No.3

以鄂马铃薯(Solanum tuberosum L.)3号为试验材料,将含有来源于黄芪的硒代半胱氨酸甲基转移酶基因(AbSMT)的质粒pBinAr进行PCR扩增以及酶切连接构建重组转化子pBI121-AbSMT,运用农杆菌转化法,设置农杆菌菌液浓度梯度、侵染时间以及共培养时间组合进行遗传转化,并且通过PCR鉴定以及GUS组织化学染色进行基因表达验证。结果表明,筛选抗生素是50.00 mg/L的Kan,农杆菌菌液浓度为0.50(OD600 nm),侵染时间为8 min,共培养时间为3 d,能够实现鄂马铃薯3号高效遗传转化,同时成功构建了AbSMT基因转化的鄂马铃薯3号。

Using Hubei potato(Solanum tuberosum L.)No. 3 as the experimental material,PCR amplification was performed on the plasmid pBinAr containing selenocysteine methyltransferase gene(AbSMT)from Astragalus membranaceous. Then,the recombinant transmitter pBI121-AbSMT was constructed by enzyme digestion and ligation. The combination of Agrobacterium solution concentration gradient,infection time,and co-culture time was set for genetic transformation,and gene expression was verified by PCR identification and GUS histochemical staining. The results showed that the selected antibiotic was 50.00 mg/L Kan. Agrobacterium solution concentration was 0.50(OD600 nm);The infection time was 8 min. Co-culture time of 3 d could realize the efficient genetic transformation of Hubei potato No. 3. At the same time,the AbSMT gene transformed Hubei potato No. 3 was successfully constructed.