Aida Ryutaro
National Institute of Floricultural Science, National Agriculture and Food Research Organization, Tsukuba, Ibaraki, Japan.
Methods Mol Biol. 2012;847:267-74. doi: 10.1007/978-1-61779-558-9_23.
This chapter describes an Agrobacterium tumefaciens-mediated transformation protocol for torenia, a plant that has several useful characteristics and is primarily used for ornamental and experimental purposes. Leaf segments of torenia were co-cultured with A. tumefaciens containing a vector plasmid for 7 days at 22°C under dark conditions on Murashige and Skoog (MS) medium containing 1 mg/L benzyladenine, 1 mg/L indoleacetic acid, and 100 μM acetosyringone. Subsequent culturing at 25°C under a 16-h photoperiod with fluorescent light on MS medium containing 1 mg/L benzyladenine, 300 mg/L carbenicillin, and selection agent (300 mg/L kanamycin or 20 mg/L hygromycin) allowed for transformant selection. Transgenic shoots were obtained from green compact calli after 2-3 months of culture in the selection medium. This method can achieve a transformation rate of approximately 5% (transformants/explant).
本章介绍了一种利用根癌农杆菌介导的蓝猪耳转化方法，蓝猪耳是一种具有多种有用特性的植物，主要用于观赏和实验目的。蓝猪耳的叶片切段在含有1 mg/L苄基腺嘌呤、1 mg/L吲哚乙酸和100 μM乙酰丁香酮的Murashige和Skoog（MS）培养基上，于22°C黑暗条件下与含有载体质粒的根癌农杆菌共培养7天。随后在含有1 mg/L苄基腺嘌呤、300 mg/L羧苄青霉素和选择剂（300 mg/L卡那霉素或20 mg/L潮霉素）的MS培养基上，于25°C、16小时荧光光照周期下进行培养，以筛选转化体。在选择培养基中培养2 - 3个月后，从绿色紧密愈伤组织中获得转基因芽。该方法可实现约5%的转化率（转化体/外植体）。
