马铃薯光诱导型茎叶特异表达启动子ST-LS1的克隆与功能分析  

Cloning and Functional Analysis of Light-inducible, and Stem and Leaf-specific Expression Promoter ST-LS1 in Potato (Solanum tuberosum L.)

作　　者：瞿韵[1,2] 张宁[2] 常璟[2] 晋昕[2] 文义凯[2] 司怀军[1,2] 王蒂[1] 

机构地区：[1]甘肃省干旱生境作物学省部共建国家重点实验室培育基地,甘肃省作物遗传改良与种质创新重点实验室,甘肃兰州730070 [2]甘肃农业大学生命科学技术学院,甘肃兰州730070

出　　处：《农业生物技术学报》2013年第7期828-837,共10页Journal of Agricultural Biotechnology

基　　金：甘肃省农业科技创新项目(No.GNCX-2011-49)

摘　　要：叶片是植物光合作用器官,在能量固定和利用中具有重要作用,研究光诱导型茎叶特异表达启动子的作用元件及其功能对于其调控基因的表达研究具有重要的理论意义和应用价值。本研究用PCR技术从马铃薯(Solanum tuberosum L.)基因组中分离了光诱导型茎叶特异表达启动子ST-LS1的1556bp序列,序列分析表明,该片段与已报道的ST-LS1启动子(Gen Bank accession No.X04753.1)有99.68%的同源性,包括参与芽的特定表达和光反应顺式作用元件as-2-box、光效应顺式作用元件G-box等。将该片段与GUS基因融合,构建了植物表达载体pBⅠ121-ST-LS1,应用根癌农杆菌(Agrobacterium tumefaciens)介导法转化烟草(Nicotiana tabacum)获得了转基因植株。用qRT-PCR和GUS活性组织染色检测转基因烟草植株,结果表明,在ST-LS1启动子驱动的GUS转基因烟草植株的叶和茎中能够检测到GUS基因的表达,根中则检测不到;对黑暗、恒温光照培养、自然光条件处理20d后的转基因植株分析表明,在黑暗处理的转基因植株中GUS基因无表达,而在自然光条件处理转基因植株的叶和茎中GUS基因的表达高于恒温光照培养的转基因植株。结果可为应用基因工程改良农作物品种提供理论和应用依据。Leaf is photosynthetic organ of plants which plays an important role in energy fixation and utilization. The study of acting elements and their functions of light-inducible, and stem and leaf-specific expression promoter have important theoretical significance and application value on research for regulation of gene expression. The 1 556 bp sequence of light-inducible, stem and leaf-specific expression promoter ST- LS1 was isolated from potato （Solanum tuberosum L.） genome by PCR assay. The result from sequence analysis showed that the fragment shared 99.68% identity with the reported ST-LS1 promoter （GenBank accession No. X04753.1）, and which contained shoot-specific expression and light responsive cis-acting element as-2-box, and cis-acting element G-box with light effect. The plant expression vector pB 1121-ST-LS1 was constructed by fusing the fragment with GUS gene, which was transferred to tobacco（Nicotiana tabacum） by Agrobacterium tumeJ＇~ciens system and obtained the transgenic tobacco plants. Both qRT-PCR and histochemical assay of GUS activity revealed that the GUS gene expression could be detected in the leaf and stem of the transgenic tobacco plants transformed with GUS gene driven by ST-LS 1 promoter, while could not be detected in root of the transgenic plant. The result from the transgenic plants under treatment for 20 d with darkness, constant temperature light, natural light culture demonstrated that there was no GUS gene expression in the transgenic plants under darkness treatment, while GUS gene expression was higher in the leaf and stem of the transgenic plants under natural light culture compared with constant temperature light condition. The results could provide theoretical and applied basis for crop improvement using genetic engineering.

关 键 词：马铃薯 ST-LSl启动子 表达载体 转基因烟草 

分 类 号：S572.032[农业科学—烟草工业]