摘要: |
【目的】利用双分子荧光互补(BiFC)技术,在烟草叶肉细胞中分析玉米ADP 葡萄糖焦磷酸化酶质体型小亚基(AGPase-bt2)与丙酮酸磷酸双激酶(PPDK1)之间的相互作用。【方法】构建326-CYCHA-AGPase-bt2和326-CYNEE-PPDK1双分子荧光表达载体,转化农杆菌EHA105,瞬时浸染烟草叶肉细胞,激光共聚焦显微镜下观察AGPase-bt2和PPDK1的相互作用。【结果】双酶切试验表明,326-CYCHA-AGPase-bt2、326-CYNEE-PPDK1载体构建正确;PCR结果证实,植物表达载体成功转化到农杆菌EHA105中;浸染烟草叶片后,AGPase-bt2和PPDK1在叶肉细胞中高效表达,出现BiFC荧光信号。【结论】AGPase-bt2和PPDK1在植物细胞内存在真实互作关系。 |
关键词: AGPase-bt2 PPDK1 双分子荧光互补技术 蛋白质互作 |
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基金项目:国家自然科学基金项目(31170731,31200611);吉林省科技厅重点科技攻关项目(20130206012NY);上海市科学技术委员会项目(10DZ2271800) |
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The interactions between maize plastidial small subunit of AGPase and glycolytic key enzyme PPDK1 |
CUI Xi-yan,WANG Kuo,ZHANG Ji-xiao,et al
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Abstract: |
【Objective】The interactions between plastidial small subunit of ADP glucose pyrophosphorylase (AGPase-bt2) and phosphate dikinase (PPDK1) of maize were investigated by bimolecular fluorescence complementation (BiFC) in tobacco mesophyll cell.【Method】First,the expression vectors of 326-CYCHA-AGPase-bt2 and 326-CYNEE-PPDK1 were constructed and transformed into Agrobacterium tumefaciens EHA105.Then,it was used to dip tobacco mesophyll cell momentarily.At last,the interactions between AGPase-bt2 and PPDK1 were observed using confocal laser scanning microscope.【Result】326-CYCHA AGPase-bt2 and 326-CYNEE-PPDK1 vectors were constructed correctly and expressed successfully.AGPase-bt2 and PPDK1 were highly expressed after dipping tobacco leaves and BiFC signals appeared.【Conclusion】The interactions between AGPase-bt2 and PPDK1 in leaf cells were proved. |
Key words: ADP glucose pyrophosphorylase phosphate dikinase bimolecular fluorescence complementation protein interaction |