泡桐遗传转化系统的建立

STUDY ON GENETIC TRANSFORMATION OF PAULOWNIA

通过农杆菌 Ｔｉ 质粒介导，将模式基因（ 新霉素磷酸转移酶＋ β葡萄糖酸苷酶基因） 导入泡桐，并对影响转化及选择再生的一系列因子进行优化，创建了泡桐的遗传转化系统．优化后的系统为：用泡桐完整叶片作外植体，侵染转化的菌液浓度 Ｏ Ｄ６００ 值控制在０ ．１ ～０ ．２ ，经２０ ｍ ｍ ｏｌ／ Ｌ Ｃａ２ ＋ 处理，共培养６ｄ ；在１０ｍ ｇ／ Ｌ 卡那霉素＋ ２５０ ｍ ｇ／ Ｌ 羧苄青霉素的选择压下培养数天，随后在仅含 Ｋａｎ的选择压下继续培养，直至抗性芽的再生．通过此遗传转化系统，获得了抗 Ｋａｎ的泡桐芽，转化频率可达８４ ％ ．组织化学法检测表明， Ｇ Ｕ Ｓ 报告基因不仅在侵染的叶片中得到短暂表达。

By the agrobacterium mediated transformation,the model genes (NPTⅡ+GUS genes)contained in the vector PBI121 was introduced into the shoots of Paulownia.Moreover,a series of factors which influenced the transformation,selection and regeneration of the plants were optimized,and the genetic transformation system of Paulownia was originally established.The optimized system was as follows.The whole leaf was used as explants.The density of agrobacterium was within OD 600 0.1～0.2.Ca 2+ treatment was applied after the explants dipped into the bacterium solution.The co culture between the explants and bacterium was for six days.The transformed callus and shoots were screened in the medium of both 10 mg/L Kan and 250 mg/L Carb and then in the medium of only Kan.By the system,the transformation efficiency was higher to 84 percent.According to the detection of histochemistry,the report gene GUS has expressed not only transiently in the leaves,but also steadily in the shoots and callus derived.