El-Shemy Hany, Khalafalla Mutasim, Wakasa Kyo, Ishimoto Masao
National Agricultural Research Center for Western Region, 6-12-1 Nishifukatsu, Fukuyama, Hiroshima, 721-8514 Japan.
Cell Mol Biol Lett. 2002;7(2B):709-19.
Two plasmid vectors were introduced into soybean (Glycine max (L.) Merr.) and azuki bean (Vigna angularis Willd. Ohwi and Ohashi) using different transformation systems. Azuki bean epicotyl explants were prepared from etiolated seedlings and co-cultivated with Agrobacterium tumefaciens for 2 days. Adventitious shoots were developed from the callus of the explants on a regeneration medium containing hygromycin, and the shoots were excised and transferred to a rooting medium containing hygromycin at the same concentration. Rooting shoots were transferred to soil and grown in a glass-house to produce viable seeds. PCR analysis confirmed clearly the presence of the hpt gene in most of the azuki beans regenerated under hygromycin selection. A soybean embryogenic suspension culture was generated from immature cotyledons, and used for the introduction of plasmids by particle bombardment. Hygromycin-resistant embryogenic clones were isolated after 8 weeks of hygromycin selection, and then the green clones were matured on the differentiation medium. After desiccation, the embryos were germinated on the rooting medium, and the plants were transferred to soil in a glass-house. More than 50% of the regenerated soybean plants tolerant to hygromycin yielded the hpt fragment on PCR analysis. The azuki bean transformants were obtained more rapidly and with higher efficiency than the soybean transformants.
利用不同的转化系统将两种质粒载体导入大豆（Glycine max (L.) Merr.）和小豆（Vigna angularis Willd. Ohwi and Ohashi）。从小豆黄化苗制备上胚轴外植体，并与根癌农杆菌共培养2天。外植体愈伤组织在含有潮霉素的再生培养基上形成不定芽，将芽切下并转移至含有相同浓度潮霉素的生根培养基上。生根的芽转移到土壤中，在温室中生长以产生可育种子。PCR分析清楚地证实，在潮霉素选择下再生的大多数小豆中存在hpt基因。从不成熟子叶产生大豆胚性悬浮培养物，并用于通过粒子轰击导入质粒。在潮霉素选择8周后分离出抗潮霉素的胚性克隆，然后将绿色克隆在分化培养基上成熟。干燥后，胚在生根培养基上萌发，植株转移到温室中的土壤中。PCR分析表明，超过50%的耐潮霉素再生大豆植株产生hpt片段。与大豆转化体相比，小豆转化体的获得更快且效率更高。
