中间锦鸡儿CiMYBJ2基因克隆及表达分析  

Cloning and Expression Analysis of Ci MYBJ2 Gene from Caragana intermedia

作　　者：邢丹丹[1] 韩晓敏[1] 冯宗琪 杨杞[1] 齐力旺[2] 刘福军 王瑞刚[1] 李国婧[1] 

机构地区：[1]内蒙古农业大学生命科学学院,呼和浩特010018 [2]中国林业科学研究院林业研究所,北京100091 [3]内蒙古赤峰市松山区安庆沟林场,赤峰024000

出　　处：《农业生物技术学报》2014年第12期1514-1524,共11页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金项目(No.31360056);教育部博士学科点基金项目(No.20121515110007);国家高技术发展研究计划863项目(No.2011AA100203)

摘　　要：MYB转录因子是植物体内最大的转录因子家族之一,参与植物生长发育的调控、对逆境的响应等生理过程。本研究通过RACE技术从中间锦鸡儿(Caragana intermedia)中克隆了一个MYB转录因子Ci MYBJ2(Gen Bank登录号:KJ937783),并对其在植物中的表达及其启动子进行了初步研究。本研究克隆得到该基因c DNA全长1 368 bp,3'UTR长196 bp,5'UTR长203 bp。其中开放阅读框长969 bp,编码一个含322个氨基酸的亲水蛋白。g DNA序列长1 285 bp,有2个内含子和3个外显子。采用实时荧光定量PCR技术分析了Ci MYBJ2的表达模式,结果发现,在高温、脱水、Na Cl、干旱等处理条件下,Ci MYBJ2基因对不同胁迫均有响应。在高温处理后,Ci MYBJ2基因的表达量在1和3 h明显降低,之后维持一个较低值,在不同时间点基因表达量与0 h相比差异均达到极显著水平(P<0.01);在脱水处理1 h后Ci MYBJ2基因的表达量上升,且与0 h相比差异极显著(P<0.01),然后逐渐降低,在3 h Ci MYBJ2基因的表达量与0 h相比差异不显著(P>0.05),在8和12 h该基因的表达量与0 h相比差异均达到极显著(P<0.01),在24 h的表达量是0 h的一半,且差异显著(P<0.05);在Na Cl处理1 h后Ci MYBJ2基因的转录水平开始增加,3 h达到最高,之后总体呈下降趋势,在8 h表达量最低,在1、3、8和24 h该基因的表达量与0 h相比差异极显著(P<0.01),在12 h Ci MYBJ2基因的表达量与0 h相比差异不显著(P>0.05);干旱处理6 d后,Ci MYBJ2基因的表达量上升,之后一直下降,且与0 d相比差异达到极显著水平(P<0.01),15 d表达量最低。研究结果暗示Ci MYBJ2基因可能在植物对胁迫的响应过程中起作用。利用染色体步移技术克隆了中间锦鸡儿Ci MYBJ2基因启动子序列,长度为873 bp,分析显示,该启动子具有响应多种非生物胁迫的顺式作用元件。研究结果为进一步研究Ci MYBJ2基因功能和表达调控提供了依据。MYB transcription factor is one of the largest family of transcription factors in plants which play impotant roles in regulation of plant growth and development, and response to many environmental stimuli. In this study, a gene named CiMYBJ2 （GenBank accession No.KJ937783） was cloned by RACE technique fromCaragana intermedia and its expression level along with the promoter was analysed. The full-length cDNA of CiMYBJ2 was 1 368 bp with the 3’UTR of 196 bp and the 5’UTR of 203 bp. The ORF of CiMYBJ2 was 969 bp, encoding a protein of 322 amino acids which was hydrophilia. The full-length gDNA was 1 285 bp, including two introns and three exons. The expression pattern of CiMYBJ2 analysed by qRT-PCR showed that it was altered by heat, dehydration, NaCl and drought treatments. After heat treatment, the expression of CiMYBJ2 decreased at 1 and 3 h, and then maintained a lower level expression. the change of gene expression at different time points was extremely significant compared with the 0 h （P〈0.01）. After dehydration treatment for 1 h, the expression of CiMYBJ2 increased shortly, and then gradually decreased at 3 h, and the expression difference was not significant compared with the 0 h （P〉0.05）. After 8 h , the expression of the gene showed extremely significant comapred with that of the 0 h （P〈0.01）, and the expression level of the gene at 24 h reached only half of that at 0 h （P〈0.05）. Under NaCl treatment, the transcript level of CiMYBJ2 began to increase after 1 h, and reached the maximum at 3 h. After that the overall trend kept downward, the expression dropped to the lowest level at 8 h, and expression of the gene was extremely significant at different time points except for 12 h （P〈0.01）. After drought treatment of 6 d, the expression of CiMYBJ2 increased slightly, and dropped significanly after compared with 0 d （P〈0.01）, reached the minimum expression at 15 d. The promoter sequence of CiMYBJ2 with 873 bp in length was obtained by genome walking technique and ma

关 键 词：中间锦鸡儿 CiMYBJ2基因 实时荧光定量PCR 启动子 

分 类 号：Q785[生物学—分子生物学]