小黑杨Rubisco小亚基基因启动子的克隆及表达活性分析  

Cloning and Expression Activity Analysis of Rubisco Small Subunit Gene Promoter in Populus×xiaohei

作　　者：曹佳玉 曹丽娜 张俏艺 张双 胡志宝 赵唐锐 许志茹[3] 李春明[1,2] 全先奎 刘关君[1,2] CAO Jiayu;CAO Lina;ZHANG Qiaoyi;ZHANG Shuang;HU Zhibao;ZHAO Tangrui;XU Zhiru;LI Chunming;QUAN Xiankui;LIU Guanjun(State Key Laboratory of Tree Genetics and Breeding,Northeast Forestry University,Harbin 150040;College of Forestry,Northeast Forestry University,Harbin 150040;College of Life Sciences,Northeast Forestry University,Harbin 150040)

机构地区：[1]林木遗传育种全国重点实验室(东北林业大学),哈尔滨150040 [2]东北林业大学林学院,哈尔滨150040 [3]东北林业大学生命科学学院,哈尔滨150040

出　　处：《植物研究》2024年第4期625-633,共9页Bulletin of Botanical Research

基　　金：黑龙江省自然科学基金联合引导项目(LH2022C015)。

摘　　要：核酮糖-1,5-二磷酸羧化酶/加氧酶(Rubisco)是光合作用中决定碳同化速率的关键酶,其中小亚基(rbcS)是由核基因编码,并且主要在叶片中表达。利用RT-qPCR技术确定了小黑杨(Populus×xiaohei)叶片中高表达的Rubisco小亚基基因PxrbcS1和PxrbcS2基因,并克隆了其上游2 240、2 174 bp的启动子。对其进行启动子元件分析结果表明,PxrbcS1和PxrbcS2的启动子具有多个与光诱导表达相关的元件,例如G-box、MRE和Box4等元件。构建pBI-121-p PxrbcS1::GUS和pBI-121-p PxrbcS2::GUS植物表达载体并遗传转化84K杨(P. alba×P. glandulosa)。GUS组织化学染色以及qPCR表达分析表明PxrbcS1和PxrbcS2的启动子能够驱动GUS基因在84K杨叶片特异性高表达。总之,上述结果表明该研究成功地从小黑杨中克隆了具有叶片高表达活性的PxrbcS1和PxrbcS2的启动子,该启动子可应用到包括杨树在内的植物光合作用相关的基因功能研究以及提高植物光合作用的遗传操作中。Ribulose-1,5-diphosphate carboxylase/oxygenase(Rubisco)is a key enzyme in photosynthesis,in which small subunits(rbcS)are encoded by nuclear genes and mainly expressed in leaves.In this study,the Rubisco small subunit genes PxrbcS1 and PxrbcS2,which are highly expressed in Populus×xiaohei’s leaves,were determined by RT-qPCR technology,and their upstream promoters of 2240 and 2174 bp were cloned respectively.The results of promoter element analysis showed that the promoters of PxrbcS1 and PxrbcS2 had multiple elements related to light induced expression,including G-box,MRE and Box4 elements.Plant expression vectors of pBI-121-pPxrbcS1::GUS and pBI-121-pPxrbcS2::GUS were constructed and genetically transformed into 84K poplar(P.alba×P.glandulosa).GUS histochemical staining and qPCR expression analysis showed that the promoters of PxrbcS1 and PxrbcS2 could drive the GUS gene to express in 84K poplar leaves with high specificity.In conclusion,the above results showed that PxrbcS1 and PxrbcS2 promoters with high leaf expression activity were successfully cloned from P.×xiaohei,and the promoters might be applied to the study of gene functions related to plant photosynthesis and genetic operations to improve photosynthesis in plants,including poplars.

关 键 词：杨树 Rubisco小亚基启动子 克隆 调控元件 遗传转化 GUS染色 

分 类 号：Q37[生物学—遗传学]