瓜叶菊‘大花’Mlo基因RNAi载体的构建及其遗传转化研究

为进一步研究瓜叶菊Mlo基因的功能以及为将来获得具有广谱与持久的白粉菌抗性瓜叶菊种质资源奠定基础,以瓜叶菊(Pericallis hybrida B.Nord)‘大花’为试验材料,克隆瓜叶菊‘大花’Mlo基因保守片段,构建了瓜叶菊‘大花’RNAi载体;同时构建了瓜叶菊‘大花’高频再生体系;并利用根癌农杆菌介导法转化瓜叶菊‘大花’进而建立了遗传转化体系。经酶切鉴定,成功构建了Mlo基因的RNAi载体质粒pTCK303-mlo-RNAi,构建了瓜叶菊‘大花’高频再生体系,最终确定瓜叶菊‘大花’最佳愈伤诱导培养基为:MS+6-BA 2.4mg/L+NAA 1.0mg/L+KT 0.3mg/L+2,4-D 1.0mg/L;瓜叶菊‘大花’愈伤组织的分化培养最适培养基为:MS+6-BA 2.0mg/L+NAA0.1mg/L;瓜叶菊‘大花’生根培养的最佳培养基为1/2MS+NAA 0.5mg·L-1,并且初步探索了瓜叶菊‘大花’Mlo基因转化体系。

Construction of RNAi vector of Mlo gene and its transformation of Pericallis hybrida B. Nord

This research aims to further study the function Mlo gene in Pericallis hybrida B. Nord and to obtain the powdery mildew resistant resources with a broad spectrum and long lasting characteristics. the Pericallis hybrida B. Nord plants with big flowers were used as materials and the conservative fragment of Mlo gene was cloned for constructing the RNAi vector and high frequency regeneration system of Pericallis hybrida B. Nord.By identifica-tion of restriction enzyme digestion,the RNAi vector (pTCK303 mlo RNAi)of gene Mlo was successfully obtained,and the regeneration system was constructed.The preferable medi-um for differentiation was MS+6 BA 2.4 mg/L+NAA 1.0 mg/L+KT 0.3 mg/L+2 ,4 D 1.0 mg/L;The best medium for regeneration of Pericallishybrida B. Nord was MS+6 BA 2.0 mg/L+NAA 0.1 mg/L;the optimum shoot-rooting medium was 1/2MS+NAA 0.5 mg/L.The transformation system of Mlo gene of Pericallis hybrida B. Nord was also initially in-vestigated.