菊基二磷酸合成酶基因：启动子腺体特异性，过表达调控除虫菊酯合成和菊科植物中该基因分子进化研究

发布时间：2021-04-19 16:10

　　除虫菊是一种重要的经济作物,以其花头富含有高量的杀虫次生代谢物除虫菊酯著称。除虫菊酯能在除虫菊地上部分合成,但是其中含量最高的合成器官为子房瘦果。前人证实位于除虫菊花和瘦果以及果皮外分泌管上的腺体组织能参与除虫菊酯的合成代谢。除虫菊酯合成途径的第一个关键步骤是利用菊基焦磷酸合成酶（CDS）催化两分子的烯丙基二磷酸（DMAPP）产生菊基二磷酸（CPP）。除虫菊TcCDS基因由FDS因家族通过基因复制进化而来,除虫菊TcCDS基因与菊科近缘植物青蒿体内的FDS基因具有70%的同源性。我们首次克隆了除虫菊TcCDS基因启动子并分析其腺体组织特异性,并且分析了启动子序列上结合不同转录因子以及响应环境和发育条件等能影响TcCDS表达和除虫菊酯合成的顺式元件,这为将来除虫菊更进一步的实验提供了一定的基础理论。同时,为利用腺体特异启动子所进行的代谢工程从而提高植物次生代谢物含量和产量提供了帮助。在以前的研究中,通过传统育种以及化学激素处理（乙烯利、矮壮素、多效唑）均能提高除虫菊酯的含量,但是相对于调控生物合成途径的代谢工程具有一定的局限性。目前还未有人尝试通过代谢工程提高除虫菊酯含量。在青蒿中,已有... 

【文章来源】：华中农业大学湖北省 211工程院校 教育部直属院校

【文章页数】：104 页

【学位级别】：博士

【文章目录】：
Abstract
摘要
List of abbreviations
1 General Introduction
    1.1 Morphology and Flower development
    1.2 History and Cultivation area
    1.3 Plant grandular trichome:specialized metabolites factories
    1.4 Chemistry of pyrethrins
    1.5 Role of pyrethrins as natural insecticide
    1.6 Pyrethrins Biosynthesis
    1.7 Trichome specific promoter
    1.8 Metabolic engineering of plant secondary metabolites
    1.9 Molecular evolution and phylogeny of the nuclear gene
    1.10 Objective of the thesis
2 Molecular Clining and Characterization of the Trichome Specific ChrysanthemylDiphosphate Synthase Promoter from Tanacetum cinerariifolium
    2.1 Abstract
    2.2 Introduction
    2.3 Materials and methods
        2.3.1 Plant materials
        2.3.2 Promoter cloning
        2.3.3 DNA sequence analysis
        2.3.4 Vector construction
        2.3.5 Plant transformation
        2.3.6 Gus assay
    2.4 Results
        2.4.1 Characterization of the promoter DNA sequence
        2.4.2 Tissue specificity of the TcCDS promoter in tobacco
    2.5 Discussion
3 Overexpression of the genes Chrysanthemyl Diphosphate Synthase （TcCDS） increasedPyrethrins content in Tanacetum cinerariifolium
    3.1 Abstract
    3.2 Introduction
    3.3 Material and Methods
        3.3.1 Plant materials
        3.3.2 Construction of pBINPLUS-CDS and transformation of Agrobacterium tumefaciens strain AGL-0
        3.3.3 Genetic transformation of T cinerariifolium
        3.3.4 Selection and regeneration of transformants
        3.3.5 DNA isolation and PCR analysis
        3.3.6 Phenotype analysis of transgenic T cinerariifolium plants
        3.3.7 RNA isolation and real-time qPCR analysis
        3.3.8 Pyrethrins Extraction methods for HPLC Analysis
        3.3.9 Pyrethrins evaluation by High-performance liquid chromatography （HPLC）
        3.3.10 Statistical analysis
    3.4 Results
    3.5 Discussion
4 Molecular evolution and phylogenetic utility of the chrysanthemyl diphosphatesynthase （TcCDS） gene,resolving relationship within Chrysanthemum
    4.1 Abstrace
    4.2 Introduction
    4.3 Materials and methods
        4.3.1 Plant material and genomic DNA isolation
        4.3.2 Nested PCR amplification and purification
        4.3.3 Cloning and sequencing
        4.3.4 DNA sequenco analysis
    4 Result
        4.1 Gene characterization and genetic analysis
        4.2 Number of nueleotide substitutions per site
        4.3 Synonymous and nonsynonymous substitutions
        4.4 Phylogenetic analysis
    4.5 Discussion
5 General Conclusion and Prospects
    5.1 Molecular Cloning and Characterization of the Trichome Speoitic Chrysanthemyl Diphosphate Promoter from Tanacetum cinerariifolium
    5.2 Overexpression of the genes Chrysanthemyl Diphosphate Synthase（TcCDS）increasedpyrethrins content in Tanacetum cinerariifolium
    5.3 The molecular evolution and phylogenetic utility of the chrysanthemyl diphosphatesynthase （CDS） gene in Chrysanthemum
References
Acknowledgements
Resume
List of Publications


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期刊论文
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[2]菊花起源的RAPD分析[J]. 戴思兰,陈俊愉,李文彬.  植物学报. 1998(11)



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