Shrawat Ashok K, Good Allen G
Department of Biological Sciences, University of Alberta, Edmonton, AB, Canada.
Methods Mol Biol. 2011;710:355-72. doi: 10.1007/978-1-61737-988-8_24.
A critical step in the development of a robust Agrobacterium tumefaciens-mediated transformation -system for cereal crop plants is the establishment of optimal conditions for efficient T-DNA delivery into target tissue, from which plants can be regenerated. Although, Agrobacterium-mediated transformation of cereals is an important method that has been widely used by many laboratories around the world, routine protocols have been established only in specific cultivars within a species and with specific tissues of high regeneration potential. Cocultivation of highly embryogenic callus tissue or healthy immature embryos with A. tumefaciens is considered one of the critical factors in successful genetic transformation of crop plants. Immature embryos collected only from vigorously growing healthy and green plants grown in the field or in the well-conditioned greenhouse are the ideal target for genetic transformation of recalcitrant crop species. Here, we describe an Agrobacterium-mediated transformation method that uses immature embryos as the starting material for inoculation with Agrobacterium. The aim of this chapter is to provide the key steps/components involved in Agrobacterium-mediated transformation of cereal crops. However, these steps or components often vary between protocols and from laboratory to laboratory, and can be optimized or modified based on the requirement of a specific cultivar or species.
对于谷物作物而言，建立一种强大的根癌农杆菌介导的转化系统的关键步骤是确立将T-DNA有效导入靶组织的最佳条件，以便能够从该靶组织再生出植株。尽管根癌农杆菌介导的谷物转化是一种重要方法，已被世界各地许多实验室广泛应用，但常规方案仅在一个物种内的特定品种以及具有高再生潜力的特定组织中得以确立。将高度胚性愈伤组织或健康的未成熟胚与根癌农杆菌共培养被认为是作物成功进行遗传转化的关键因素之一。仅从田间或条件良好的温室中茁壮生长的健康绿色植株上采集的未成熟胚是难转化作物物种进行遗传转化的理想靶标。在此，我们描述一种根癌农杆菌介导的转化方法，该方法使用未成熟胚作为接种根癌农杆菌的起始材料。本章的目的是提供根癌农杆菌介导的谷物作物转化所涉及的关键步骤/组成部分。然而，这些步骤或组成部分在不同方案之间以及不同实验室之间常常有所不同，并且可以根据特定品种或物种的需求进行优化或修改。
