基因枪转化法在棉纤维细胞中瞬时表达外源基因的研究  

Study on Transient Expression of Target Gene in the Epidermal Cells of Cotton Ovules via Particle Bombardment

作　　者：于晓玲[1] 崔百明[1] 卫海滨[1] 赵平娟[1] 彭明[1] 

机构地区：[1]中国热带农业科学院热带生物技术研究所

出　　处：《棉花学报》2007年第6期419-423,共5页Cotton Science

基　　金：国家重点基础研究发展计划(973):2004CB117307

摘　　要：试验运用了0 d,1 d的棉胚珠为受体,GUS基因为报告基因,用基因枪直接对胚珠表皮细胞进行轰击。整个表达载体包括两个部分,第一部分是由CaMV35S调控的GUS基因,第二部分是由棉纤维特异表达启动子(RDL1)调控的目标基因,因此目标基因与GUS报告基因能在同一个单纤维细胞中同时表达,从而提供了一个直观的在单纤维细胞中验证目标基因的表达与纤维发育关系的方法。研究中对受体棉花胚珠的发育时间、基因枪的轰击压力、轰击距离及轰击次数等影响转化效率的因素进行了分析优化。结果表明,最佳的转化组合条件是:以开花一天的棉花胚珠为受体,当轰击压力为1100 psi,轰击距离为9cm,轰击次数为2次时可达到最高的转化率,并能观察到稳定的GUS表达。Cotton fibers are unicellular trichomes which differentiate from epidermal cells of developing cotton ovules. They have been considered as an ideal model for studying plant cell elongation and cell wall biogenesis. A large number of regulatory genes are involved in the process of fiber differentiation and development. Only a few is known how they control and regulate the entire process of fiber devel opment. Because generating transgenic cotton needs a long period of time and transformation efficiency of cotton is still low, it is necessary to develop a fast and efficient transient expression assay system for characterization of cotton fiber-specific expression genes and promoters as well as test of their function. Cotton ovule culture （0-1 DPA） and β-glucuronidase gene （GUS） were used in biolistic transformation experiment. The transient expression vector contains two expression cassettes. GUS reporter gene is driven by CaMV35S promoter, the target gene is under control of cotton fiber-specific promoter RDL1. Because GUS reporter gene and target gene simultaneously express in single fiber cell, the transient expression assay system could provide an effective method to investigate the rela tionship between the expression of target gene and the development process of fiber in transformed single cotton fiber cell. The conditions for bombardment were optimized to enhance the transformationefficiency . The result showed that the highest transformation efficiency of GUS was obstained when one day postanthesis （DPA） ovules were applied. The optimal condition for bombardment include he-lium pressures of 1100 psi, distance of 9 cm and bomb times of twice.

关 键 词：棉纤维 基因枪 瞬时表达 GUS 

分 类 号：S562[农业科学—作物学]