基因枪及其与农杆菌相结合的茶树外源基因转化条件优化

doi:10.13305/j.cnki.jts.2005.04.004

摘要/Abstract

摘要： 以GUS为报告基因，对基因枪介导的转化茶树愈伤的有关参数进行优化。比较了不同的预培养条件，含有PVP的培养基可提高转化频率。渗透处理不仅不能提高GUS的表达率，反而使愈伤再生困难。每枪DNA和钨粉的用量分别为0.25 μg，125 μg时，可得到较高的转化率。在轰击压力7 MPa，射程5 cm的条件下轰击一次，不论对GUS表达还是愈伤的再生都是较为适合的轰击条件。比较了农杆菌法（AGR），基因枪法(BOM)及农杆菌与基因枪结合(BTA，BPA和BOA)等方法对茶树遗传转化效率的影响。瞬间表达的结果及抗性愈伤筛选的结果都显示，农杆菌与基因枪结合使用比两种方法单独使用更有助于提高茶树转化的效率。

关键词: 茶树, 遗传转化, 基因枪, 农杆菌

Abstract: A microprojection protocol using particle delivery system and GUS report gene was developed for tea plant (Camellia sinensis) callus. Comparison of several pre-bombardment culture media showed the highest transient gene expression in the media with PVP. Instead of making contribution to improving GUS expression, osmotic pre-treatment would cause the target tissue having difficulty in regeneration. High transformation rate was obtained using 125 μg tungsten coated with 0.25 μg DNA per shot. Bombarding tea callus at 7 MPa with a target distance of 5 cm and one time was suitable for both GUS activity and regeneration. Several transformation methods, including Agrobacterium-mediated transformation(AGR), particle bombardment (BOM), and combination of the two methods (BTA, BPA and BOA), have been tested and compared with each other. Based on the results of GUS expression and survival rates of the resistant calli, the method combining Agrobacterium and particle bombardment systems is suitable to apply for tea plant transfortion.

Key words: Tea plant (Camellia sinensis), Genetic transformation, Particle bombardment, Agrobacterium tumefaciens

吴姗, 梁月荣, 陆建良, 黎昊雁. 基因枪及其与农杆菌相结合的茶树外源基因转化条件优化[J]. 茶叶科学, 2005, 25(4): 255-264. doi: 10.13305/j.cnki.jts.2005.04.004.

WU Shan, LIANG Yue-rong, LU Jian-liang, LI Hao-yan. Combination of Particle Bombardment-mediated and Agrobacterium-mediated Transformation Methods in Tea Plant[J]. Journal of Tea Science, 2005, 25(4): 255-264. doi: 10.13305/j.cnki.jts.2005.04.004.

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