小立碗藓PpAGO7基因启动子分离及其启动活性分析  

Separation and Functional Analysis of the Promoter of PpAGO7 Gene from Physcomitrella patens

作　　者：高运通 潘沈元[1] 王爱民[1] 

机构地区：[1]江苏师范大学生命科学学院,江苏徐州221116

出　　处：《西北植物学报》2015年第3期465-471,共7页Acta Botanica Boreali-Occidentalia Sinica

基　　金：科技部转基因生物新品种培育科技重大专项子课题(2014ZX08012-002)

摘　　要：AGO7蛋白在多种植物中被发现并预测在叶片生长发育过程中起作用,但是在高等植物中最古老且唯一没有维管束的苔藓植物中却未有报道。该文通过BLAST比对水稻OsAGO7基因编码的氨基酸序列得到小立碗藓AGO7蛋白编码基因PpAGO7,扩增PpAGO7基因起始密码子上游的启动子序列,利用在线软件PlantCARE和PLACE分析该启动子的结构特征;并构建启动子分析载体pPpAGO7-GUS,转化拟南芥,通过转基因拟南芥GUS染色结果分析推测PpAGO7启动子的启动特性。结果显示:(1)PpAGO7基因启动子序列中含有大量的光反应有关的顺式作用元件以及数个分生组织相关和防御与胁迫相关作用元件。(2)T2代转基因拟南芥的GUS染色结果表明,PpAGO7基因启动子会启动GUS在拟南芥的不同部位和不同生长时期表达,而且在根尖、叶片顶端、雄蕊的花药、雌蕊的柱头和种子等部位的染色较其他部位更深。(3)生长在光照强度1 000lx和4 000lx下转基因拟南芥的GUS酶活性比光照强度7 000lx和10 000lx下的低。研究表明所克隆的PpAGO7基因启动子具有组成性启动活性,且在生长旺盛的部位启动活性较强,此外其启动活性还受到光照因素的影响,为进一步研究小立碗藓的PpAGO7基因功能提供了重要依据。The AGO7 was found in many plants and was predicted to play a role in the process during growth and development of leaves in higher plants.However,it was not reported in bryophytes which was one of the oldest and the only one which had no vascular bundle in higher plants.By comparison to the amino acid sequence of OsAGO7 in rice with BLAST tool,we obtained the AGO7 protein code gene in Physcomitrella patens named PpAGO7.We also cloned the promoter sequence of PpAGO7 and analyzed its sequence characters by using online software Plant CARE and PLACE.Then the promoter analysis vector pPpAGO7-GUS was built and transferred into Arabidopsis to detect the initiation activity.The result indicated that：（1）The promoter sequence contained large amount of cis-acting elements related to the photo reaction and a plurality of elements related to the meristem and defense and oppression related elements.（2）According to the staining result of the T2 transgenic plants,GUS staining was found in different tissues andduring different growth periods.Moreover,the staining color was much deeper in the root,leaf tip,anther of stamen,stigma of pistil and seeds than in other tissues.（3）The GUS enzymatic activity of the transgenic Arabidopsis under the light intensity condition 1 000 lx or 4 000 lx was lower than that under the light intensity condition 7 000 lx or 10 000 lx.All results indicted that the cloned PpAGO7 promoter had constitutively promoting activity and was influenced by light intensity.

关 键 词：小立碗藓 PpAGO7 启动子 

分 类 号：Q785[生物学—分子生物学] Q789