Vain Philippe, Thole Vera
Department of Crop Genetics, John Innes Centre, Norwich, Colney, UK.
Methods Mol Biol. 2009;478:203-26. doi: 10.1007/978-1-59745-379-0_13.
During the past 25 years, the molecular analysis of transgene insertion patterns and sites in plants has greatly contributed to our understanding of the mechanisms underlying transgene integration, expression, and stability in the nuclear genome. Molecular characterization is also an essential step in the safety assessment of genetically modified crops. This chapter describes the standard experimental procedures used to analyze transgene insertion patterns and loci in cereals and grasses transformed using Agrobacterium tumefaciens or direct transfer of DNA. Methods and protocols enabling the determination of the number and configuration of transgenic loci via a combination of inheritance studies, polymerase chain reaction, and Southern analyses are presented. The complete characterization of transgenic inserts in plants is, however, a holistic process relying on a wide variety of experimental approaches. In this chapter, these additional approaches are not detailed but references to relevant bibliographic records are provided.
在过去25年里，对植物中转基因插入模式和位点的分子分析极大地促进了我们对转基因整合、表达及在核基因组中稳定性的潜在机制的理解。分子特征分析也是转基因作物安全性评估的关键步骤。本章介绍了用于分析通过根癌农杆菌介导或DNA直接转移转化的谷物和禾本科植物中转基因插入模式和位点的标准实验程序。文中展示了通过遗传研究、聚合酶链反应和Southern分析相结合来确定转基因位点数量和构型的方法及方案。然而，对植物中转基因插入片段的完整特征分析是一个依赖多种实验方法的整体过程。本章未详细阐述这些额外方法，但提供了相关文献记录的参考文献。
