烟草NtGSH2和NtPCS1基因的分子特征及其对Cd和Zn的响应

发布时间：2018-10-24 15:57

【摘要】：高等植物主要通过络合作用和区室化作用进行重金属解毒,而谷胱甘肽合酶和植物络合素合酶是植物体重金属解毒途径的关键酶。本研究以烟草(Nicotiana tabacum)为材料,克隆烟草谷胱甘肽合酶基因(NtGSH2)和植物络合素合酶基因(NtPCS1)。通过序列比对,组织表达模式,重金属诱导表达等方面分析NtGSH2和NtPCS1基因的基本分子特征。并通过异源表达和过量表达分析NtPCS1基因的生物学功能。结果显示:(1)来自不同物种的GSH2蛋白的氨基酸序列高度保守,而PCS蛋白的N端结构域序列高度保守。(2)NtGSH2和NtPCS1基因在烟草的根、茎、叶、花和种子均表达,且均在花器官中的表达量最高。(3)NtGSH2和NtPCS1基因的转录表达均受到Cd~(2+)和Zn~(2+)的诱导而上调。(4)NtPCS1基因编码植物络合素合酶蛋白,且Ag~(2+)和Cd~(2+)能够显著增加植物络合素合酶蛋白的活性。(5)NtPCS1基因的异源表达能够增强拟南芥Atpcs1突变体对Cd~(2+)的耐受性(6)NtPCS1基因的过量表达导致烟草对Cd~(2+)超敏感。(7)NtPCS1基因的过量表达导致烟草体内PCs含量的显著增加。(8)外源GSH的添加能够减弱过表达转基因烟草对Cd~(2+)的超敏感。(9)外源GSH的添加导致过表达转基因烟草体内积累更多的Cd~(2+)和Zn~(2+)。
[Abstract]:High plants detoxify heavy metals mainly by complexation and compartmentalization, and glutathione synthase and plant complexin synthase are the key enzymes in detoxification pathway of heavy metals in plants. In this study, tobacco glutathione synthase (NtGSH2) gene and plant complex synthase gene (NtPCS1) were cloned from tobacco (Nicotiana tabacum). The basic molecular characteristics of NtGSH2 and NtPCS1 genes were analyzed by sequence alignment, tissue expression patterns and heavy metal induced expression. The biological function of NtPCS1 gene was analyzed by heterologous expression and overexpression. The results showed that: (1) the amino acid sequence of GSH2 protein from different species was highly conserved, while the N-terminal domain of PCS protein was highly conserved. (2) NtGSH2 and NtPCS1 genes were expressed in roots, stems, leaves, flowers and seeds of tobacco. (3) the transcription expression of NtGSH2 and NtPCS1 genes were up-regulated by Cd~ (2) and Zn~ (2). (4) NtPCS1 gene encodes plant complex synthase protein. Ag~ (2) and Cd~ (2) could significantly increase the activity of plant complexin synthase protein. (5) the heterologous expression of NtPCS1 gene could enhance the tolerance of Arabidopsis Atpcs1 mutants to Cd~ (2). (6) the overexpression of NtPCS1 gene resulted in tobacco being hypersensitive to Cd~ (2) NtPCS1 gene. (8) the addition of exogenous GSH could weaken the hypersensitivity of overexpression transgenic tobacco to Cd~ (2). (9) the addition of exogenous GSH resulted in the accumulation of more Cd~ (2) and Zn~ (2) in over-expressed transgenic tobacco.
【学位授予单位】：西南科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2

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