苹果MdCBL3基因的克隆与功能鉴定

从‘嘎拉’苹果(Malus×domestica Borkh.)中克隆MdCBL3(序列号:MDP0000155124),长852 bp。系统进化树分析表明,苹果MdCBL3与白梨亲缘关系最近。利用PlantCare数据库进行启动子顺式作用元件预测分析,MdCBL3启动子序列中存在干旱、低温、光、生长素、防御等响应元件。构建MdCBL3表达载体并利用农杆菌介导法侵染苹果愈伤组织和拟南芥。半定量RT-PCR证明MdCBL3在转基因愈伤组织中过量表达,在拟南芥中得到异源表达。表型分析发现,在盐胁迫中,与野生型相比,转基因愈伤组织和拟南芥成龄苗生长更好,盐胁迫的抗性更强。在拟南芥中异源表达MdCBL3,能提高拟南芥对干旱和低温的抗性。

Molecular Cloning and Functional Characterization of MdCBL3 in Apple

A gene named MdCBL3（MDP0000155124）was cloned from‘Gala’apple（Malus × domestica Borkh.）. The length of MdCBL3 was 852 bp. A phylogenetic tree analysis indicated that the Pyrus bretschneideri exhibited the highest sequence similarity to MdCBL3. The promoter’s cis-acting elements were analyzed and predicted using PlantCare Databases，analysis showed that the MdCBL3 promoter sequence included drought，low temperature，light，auxin，defense responsiveness etc. Construct expression vector of MdCBL3 and use Agrobacterium mediated method to infect apple callus and Arabidopsis seed. Semi RT-PCR indicated that MdCBL3 was genetically transformed into apple callus and Arabidopsis. Phenotypic analysis found that transgenic callus and Arabidopsis seedling grow better and have stronger resistance under NaCl stress compared with contrast. Demonstrated that overexpression of MdCBL3，heterogenous expression of MdCBL3 remarkably increased the tolerance of transgenic apple callus and Arabidopsis to high salinity. The study found that heterogenous expression of MdCBL3 in Arabidopsis enhenced the tolerance of drought and low temperature.