茶树农杆菌转化系统和基因枪转化系统的优化  

Optimization of Agrobacterium-mediated and Particle Bombardment-mediated Transformation Systems in Tea Plant (Camellia sinensis)

作　　者：吴姗[1] 梁月荣[1] 陆建良[1] 金惠淑[1] 吴颖[1] 

机构地区：[1]浙江大学茶叶研究所,浙江杭州310029

出　　处：《茶叶科学》2003年第1期6-10,共5页Journal of Tea Science

基　　金：国家自然科学基金(30170591);国家博士点基金(200033539)资助项目

摘　　要：在用农杆菌浸染前,将茶树外植体预培养在含有PVP(polyvinylpyrrolidone,16g/L)的预培养基上2-3d可提高转化频率.优化后的茶树基因枪转化体系,即制弹程序:60 mg/1ml钨粉悬浮液10 μl中加入1.6 μl质粒DNA(1 μg/μl),再分别加入0.1 mol/L的亚精胺4 μl,2.5 mol/L 的CaCl2 15 μl,最后定容至48 μl;每次轰击上样量为8-10 μl.基因枪转化后抗性筛选2个月,抗性愈伤组织存活率为5.0%～12.1%.When tea explants were pre-cultured on medium containing PVP 16g/L for 2-3d before infiltrated by Agrobacterium tumefaciens the transformation frequency was increased. Optimum conditions for preparing bombardment micro-particle were as follows: 10 l tungsten suspension (60 mg/1ml), 1.6 l plasmid DNA (1 g/ l), 4 l spermidin (0.1 mol/L) and 15 l CaCl2 (l2.5 mol/L). Ethanol was used to dilute the final volume up to 48 l. The loading volume for each bombardment was 8-10 l. The survival rate of resistant calli was 5.0%-12.1%, when the bombarded explants were cultured on selection media with kanamycin or hygromycin.

关 键 词：茶树 农杆菌转化系统 基因枪转化系统 优化 预培养处理 

分 类 号：S571.1[农业科学—茶叶生产加工] S188[农业科学—作物学]