拟南芥rd29A的功能鉴定及植物表达载体的构建  

Identification of promoter rd29A from Arabidopsis thaliana and the construction of plant expression vector

作　　者：吴杨[1,2] 贺俐[2] 赵书环[1] 张木清[1] 

机构地区：[1]福建农林大学农业部甘蔗生理生态与遗传改良重点实验室,福建福州350002 [2]井冈山大学生命科学学院,江西吉安343009

出　　处：《福建农林大学学报（自然科学版）》2008年第6期614-619,共6页Journal of Fujian Agriculture and Forestry University:Natural Science Edition

基　　金：国家“863”计划课题“糖料新品种选育”(2001AA241191);国家自然科学基金(30370901);教育部高等学校博士学科点专项科研基金项目(20040389009)

摘　　要：利用PCR技术从拟南芥(Arabidopsis thaliana)基因组中分离了rd29A基因上游420 bp的调控序列,序列分析表明,该片段与已报道的rd29A启动子有100%的同源性,包括DRE、ABRE等顺式作用元件.用rd29A启动子构建了具有绿色荧光蛋白(GFP)基因的植物表达载体,通过基因枪导入法转化甘蔗愈伤组织,经过抗性筛选获得再生植株,在PEG胁迫下,用荧光显微镜观察到s65t(GFP)基因在愈伤组织和叶片中表达,通过PCR鉴定获得了5株转基因植株.同时本文还构建了由rd29A调控的DREB2B基因的植物表达载体rd29A-dreb-hyg.The 420 bp upstream regulatory region of rd29A gene from Arabidopsis thaliana genome was amplified by PCR technique. Sequence analysis showed that it shared 100% identity with the reported rd29A promoter and contained several cis-acting elements including dehydration responsive elements （DRE） , ABA responsive elements （ABRE） and so on. The plant expression vector Prd- s65t was constructed with this fragment linked up with green fluorescent protein （GFP） gene controlled by rd29A promoter,which was transferred to sugarcane callus by bombardment microprojectiles. Regenerated plants were obtained by resistance selection. The expressions of s65t in the transformed sugarcane callus and leaves treated with 6 g·L^-1 PEG and 40% PEG, respectively, were detected by fluorescent microscope. Through PCR analysis, 5 plants were identified to be the transgenic. At the same time, plant expression vector rd29A-dreb-hyg was constructed with this fragment linked up with DREB2B gene controlled by inducible promoter rd29A, which will lay the material foundation for sugarcane drought-resistant transgenic breeding.

关 键 词：RD29A 启动子 甘蔗 表达载体 DREB 

分 类 号：Q786[生物学—分子生物学]