棉花GhMADS29启动子克隆及表达分析课件.pdfVIP

花 学 报 Cotton Science 20 13 25 渊4 冤院309~315 棉花 29 启动子克隆及表达分析 张文香 范术丽 宋美珍 庞朝友 魏恒玲 喻树迅* 中国农业科学院棉花研究所袁河南安阳455000 冤 摘要院以实验室克隆的 29 GeneBank 登录号院JQ682642 冤基因的cDNA 序列Blast 搜索雷蒙德氏棉的 基因组序列袁根据Blast 结果设计引物袁克隆到起始密码子上游-19 位开始的1316 bp 的序列曰利用PlantCARE 启动子在线分析软件预测其含有核心启动子元件TATA-box 和CAAT-box 袁并含有光尧温尧赤霉素尧水杨酸尧生 长素等的响应元件遥 通过替换pBI121 载体上的CaMV35S 启动子构建 29 启动子与 基因的 融合表达载体并转化拟南芥袁组织化学染色分析发现其在14 d 幼苗的根和叶 都有表达袁在萼片尧花瓣尧雌蕊尧 果瓣 也表达袁而在雄蕊和种子 不表达遥 综上所述袁我们推测 29 可能与各种开花途径有关袁与萼 片尧花瓣尧雌蕊等花器官的发育有关袁还可能与果实是否开裂有关遥 关键词院启动子曰GUS 染色曰瞬时表达曰表达分析 中图分类号院S562.035.3 文献标志码院A 文章编号院1002-7807(2013)04-0309-07 ZHANG Wen-xiang, FAN Shu-li, SONG Mei-zhen, PANG Chao-you, WEI Heng-ling, YU Shu-xun* ( 455000, ) We used the cDNA sequence of 29 as a search query to BLAST the genome sequences of L. We designed the primers to clone the promoter according to the BLAST results. We obtained a 1316-bp sequence up- stream from the -19 site of the start codon. Sequence analysis showed that this promoter region contained a TATA-box core ele- ment, a CAAT-box, and other -acting elements, including an AuxRR core, a TCA element, TC-rich repeats, an HSE, Box I, a GARE motif, and a TGA element; these elements are involved in auxin, salicylic acid, defense and stress, heat, light, and gib- berellin responses, respectively. We replaced the CaMV 35S promoter in the 2 1 vector with the 1316-bp promoter sequence to construct a fusion vector containing the 29 promoter connected to the gene. The fusion vector was designated as 121. GUS histochemical staining analysis of transgenic carrying the pSVP 121 vector showed that the pro- moter was active in the roots and leaves of 14-day-old