板栗IFRs启动子分离及表达载体构建  

Seperation and Expression Vector Construction for IFRs Promoter in Castanea mollissima

作　　者：李琳玲 陈小玲 袁红慧 程水源[1,3] 程华 

机构地区：[1]经济林木种质改良与资源综合利用湖北省重点实验室,湖北黄冈438000 [2]大别山特色资源开发湖北省协同创新中心,湖北黄冈438000 [3]武汉轻工大学生物与制药工程学院,湖北武汉430023

出　　处：《北方园艺》2017年第11期107-114,共8页Northern Horticulture

基　　金：国家自然科学基金资助项目(31600529);湖北省教育厅青年基金资助项目(Q20132909);湖北省教育厅优秀中青年科技创新团队资助项目(T20619);黄冈师范学院科技创新团队资助项目(T2016003)

摘　　要：以湖北省罗田板栗品种乌壳栗为试材,采用染色体步移的方法获得了板栗IFR启动子序列,研究了板栗黄酮代谢途径中关键基因IFR的启动子顺式作用原件及可能对雄花黄酮类物质合成的影响,并构建pCAMBIA1304融合载体。以期揭示板栗雄花黄酮代谢合成途径,IFRs启动子上游所包含的顺式元件对环境和栽培措施的响应及对板栗雄花发育的影响。结果表明:长度为1 688bp的板栗IFRs启动子序列包含多种类型的顺式作用元件,包括光响应元件GT-1motif、ASF-1motif等;与赤霉素,脱落酸、乙烯等相关的激素响应元件,如WRKY motif(赤霉素信号途径转录因子)、GCC-box(乙烯应答元件)以及Dc3(脱落酸响应元件)等;逆境胁迫相关的功能元件,如与抗损伤相关的ERF3、抗病相关的TGTCA-box等。以此推测,CmIFR基因的表达可能会受到以上光、激素、各种生物或者非生物胁迫等因素的影响。为了后期进一步研究启动子的功能,按照启动子功能区域,设计了CmIFR启动子不同长度的6个功能片段,并将这些片段替换pCAMBIA1304中的35S启动子,成功构建了由CmIFRPs启动子不同区域驱动的gus基因的植物表达载体pCAMBIA1304+CmIFRPs。The chestnut ‘Wukeli’ was used as the test material,the promoter sequence of IFR isolated from chestnut DNA according to genewalking technology.The regulation of IFR promoter and its effection in flavonoids in male flowers analyzed with cis-element informatics database.For further study,the promoter,which seperated into six fragments in different function also ligated to the expression vector pCAMBIA1304.It would help to reveal the synthesis pathway of flavonoids in chestnut flower,the cis-element function analysis also reflect the different stage and environment in female flower development.The results showed that,1 688 bp promoter sequence was isolated from the DNA of ‘Wukeli’.IFRs promoter contained many important types of cis-acting elements,hormone responsive elements,such as WRKY-motif（GA signaling pathway transcription factor）,GCC-box （ethylene response element）,and Dc3 （off acid response element）.There were also stress resistance elements as flollows,the disease resistance element TGTCA sequence,the wounding responsive element ERF3（TGACY） and so on.The prediction showed that the expression of CmIFR gene might be affected by the above factors,such as light,hormone,biological or abiotic stress.Therefore,in advance to study these promoter,the direction of the research in the follow-up function verification.Six 5′-deletions expression vectors were constructed,35S promoter in pCAMBIA1304 was replaced by these fragments.And the plant expression vector pCAMBIA1304＋CmIFRPs of gus gene driven by CmIFRPs was successfully constructed.

关 键 词：板栗 黄酮 IFR 调控元件 表达载体构建 

分 类 号：S664.203.6[农业科学—果树学]