转基因玉米的分子特征表征

发布时间：2021-12-31 08:05

　　转基因（GENETICALLY MODIFIED,GM）作物的分子特征（包括插入位点和旁侧序列等）是GM作物安全评估和监管的基本组成部分,其表征必须以个案的方式予以进行。现有的各种基于聚合酶链反应（POLYMERASE CHAIN REACTION,PCR）的GM作物分子特征的表征方法都具有独特的优缺点,任何一个单独的PCR方法几乎都不可能完全揭示一个给定GM作物的分子特征,特别是碰到宿主基因组复杂或有非预期插入的GM事件。最近开发的新一代测序技术（NEW GENERATION SEQUENCING,NGS）为GM作物分子特征的表征提供了一种潜在的替代方案,其分辨率远高于基于PCR的方法。NGS已成功应用于GM大豆和GM水稻等作物的分子特征表征,但由于玉米基因组的复杂,NGS在GM玉米分子特征表征方面的应用很少。在本研究中,我们将基于PCR和基于NGS的方法结合起来,试图揭示我国自主研发的两个GM玉米事件的分子特征（主要是插入位点和旁侧序列）。这两个GM玉米分别是IE09S034和12-5,前者是抗虫GM玉米,后者是耐除草剂和抗虫双效GM玉米。所用的PCR方法包括热不对称交错PCR（T... 

【文章来源】：上海交通大学上海市 211工程院校 985工程院校 教育部直属院校

【文章页数】：110 页

【学位级别】：硕士

【文章目录】：
摘要
Abstract
List of Abbreviations
Chapter 1 Introduction
    1.1 Worldwide Status and Division of Commercial Transgenic Crops along with its Economic Background
    1.2 Agricultural Biotechnology in China
        1.2.1 Overview of Agricultural Biotechnology in China
        1.2.2 Challenges Towards Commercialization of GM Crops in China
        1.2.3 Risk Assessment of Biotech Crops in China
        1.2.4 Labelling Policies of Biotech Crops in China
        1.2.5 GM Products in China and Scientist Perspective
    1.3 Past and Present of the Detection Methods for GM Crops and Food
        1.3.1 Introduction to Molecular Characterization of GM
        1.3.2 Methods for the Molecular Characterization of Gmo’s
        1.3.3 DNA based Methods for Identification of Unknown Flanking Sequence and Insertion Site
        1.3.4 Protein Based Detection Methods
        1.3.5 PCR Based Molecular Characterization Methods
        1.3.6 Next Generation Sequencing
    1.4 Objectives
Chapter 2 Materials and Methods
    2.1 Materials
        2.1.1 IE09S034
        2.1.2 12-5
    2.2 Methods
        2.2.1 Dna Extraction
        2.2.2 Reagents and Equipment
        2.2.3 Tail-PCR
        2.2.4 Inverse Polymerase Chain Reaction
        2.2.5 Digital PCR
        2.2.6 Southern Blot
        2.2.7 Next Generations Sequencing
Chapter 3 Results
    3.1 Result of IE09S034 event
        3.1.1 Validation of full-length of the insertion
        3.1.2 Southern blot results
        3.1.3 Digital pcr results
        3.1.4 Insertion site and flanking sequence identified by tail-pcr
        3.1.5 Insertion site and flanking sequence identified by i-pcr
        3.1.6 Insertion site and flanking sequence identified by NGS
    3.2 Results of12-5 event
        3.2.1 VALIDATION OF CRY1AB/CRY2AJ AND G10-EVO-EPSPS IN 12-5
        3.2.2 Validation of sequence between CAMV35S terminator and CAMV35S/Ubiquitin mixed promotor
        3.2.3 Validation of sequence between CAMV35S/Ubiquitin mixed promotor and PEPC
        3.2.4 Validation of sequence between PEPC and CRY1AB/CRY2AJ
        3.2.5 Validation of sequence between CRY1AB/CRY2AJ and maize Ubiquitin promotor pZmUbi-1
        3.2.6 Southern blot results
        3.2.7 Digital pcr results
        3.2.8 Insertion site and flanking sequence identified by tail-pcr
        3.2.9 Insertion site and flanking sequence identified by I-pcr
        3.2.10 Insertion site and flanking sequence identified by NGS
Chapter 4 Discussion and Conclusion
Acknowledgements
References
Publications
附录


【参考文献】：
期刊论文
[1]Global challenges of food safety for China[J]. Joseph J.JEN.  Frontiers of Agricultural Science and Engineering. 2018(03)



本文编号：3559908