去甲基化酶基因AtROS1化学诱导表达载体的构建及其在烟草中的瞬时表达  

Construction of a chemical-inducible expression vector of AtROS1 and its transient expression in tobacco

作　　者：常英英[1] 梁立雄 高亚南[1] 王颜波 丁昌俊[1] 苏晓华[1] 张冰玉[1] 

机构地区：[1]中国林业科学研究院林业研究所/林木遗传育种国家重点实验室/国家林业局林木培育重点实验室,北京100091

出　　处：《浙江农业学报》2016年第5期717-723,共7页Acta Agriculturae Zhejiangensis

基　　金：国家高技术研究发展计划(863计划)(2013AA102703);林木遗传育种国家重点实验室基本科研业务费专项资金(TGB2013010)

摘　　要：植物去甲基化酶ROS1是表观遗传中一种重要的作用因子,参与基因的表达调控,与植物的生长发育及各种逆境响应过程密切相关。以拟南芥AtROS1基因为目的基因,采用"酶切—连接"的方法构建了以17-β-雌二醇为诱导剂的植物表达载体p ER8-ROS1。将其转入农杆菌LBA4404菌株中,通过烟草瞬时表达技术,对该载体的诱导表达特性进行了研究。q PCR结果表明:17-β-雌二醇处理能够有效调控p ER8-ROS1中目的基因的表达,17-β-雌二醇最佳诱导浓度为50~100μmol·L^(-1);随着诱导时间的延长,目的基因表达量逐渐升高,在12 h后达到最高。该诱导表达载体的构建为研究植物抗逆胁迫过程中的表观遗传机制奠定了基础。Plant demethylase ROS1 was an important factor in epigenetic regulation. It could activate the expression of genes and was closely related to the processes of plant development and various stress responses. In this study,AtROS1 gene from Arabidopsis thaliana was used as the target gene,and ‘digestion-ligation＇ method was applied for constructing plant expression vector p ER8-ROS1 which could be induced by 17-β-estradiol. Then the vector was transferred to Agrobacterium tumefaciens LBA4404,and the inducible expression characteristics were verified by the transient expression system of Nicotiana tabacum. The results of q PCR showed that 17-β-estradiol could effectively induce the expression of the target gene in p ER8-ROS1 and the optimal concentration of 17-β-estradiol was 50 to 100μmol·L^（-1）. The expression level of ROS1 gene gradually increased and reached the highest level at 12 h. The construction of p ER8-ROS1 laid a good foundation for the epigenetic mechanism study in plant environmental stress.

关 键 词：pER8-ROS1 17-Β-雌二醇 化学诱导表达载体 瞬时表达 

分 类 号：Q943.2[生物学—植物学]