木薯MeTPS9基因克隆及表达特性分析

Clone and Expression Characteristics of MeTPS9 Gene in Cassava

旨在揭示木薯Me TPS9基因在干旱、低温、遮荫等非生物胁迫应答中的作用。用RT-PCR的方法从木薯叶片中克隆Me TPS9基因(Gen Bank登录号:MF276889),用MEGA软件构建系统进化树,用Dna SP软件分析基因结构变异,用Plant CARE分析启动子元件,用荧光定量PCR技术分析Me TPS9在不同组织中以及响应不同非生物胁迫的表达特性。结果显示,从木薯中克隆了一个TPS基因Me TPS9。该基因具有2 598 bp的开放阅读框,编码865个氨基酸,含有TPS家族保守结构域。系统进化树分析表明,Me TPS9与荠菜花和拟南芥中同源基因的亲缘关系较近,序列相似性分别为76.2%和75.6%。基因结构变异发现,木薯野生种和栽培种之间共有66个突变位点,其中11个为错义突变。启动子元件分析表明,Me TPS9含有干旱相关元件MBS、低温相关元件LTR、热胁迫相关元件HSE、以及ABA相关元件ABRE。实时荧光定量PCR分析表明,Me TPS9的表达量在须根中最高,在叶片中最低。而且,Me TPS9基因的表达能被干旱、低温、和遮荫处理显著诱导。Me TPS9在转录水平对木薯干旱、低温、和遮荫胁迫起调控作用,可作为候选基因进一步研究其在木薯抗逆中的作用。

This work aims to reveal the function of Me TPS9 gene in abiotic stresses（e.g.,drought,cold,and shade）in cassava.RT-PCR method was applied to clone Me TPS9 gene（Gen Bank accession number ：MF276889）from cassava leaves,MEGA software toconstruct its phylogenetic tree,Dna SP software to analysze its structural variation,Plant CARE software to analyze its promoter cis-element,and quantitative RT-PCR（q RT-PCR）to explore its expression patterns in different tissues and in response to different abiotic stresses. Resultsshowed that a TPS（trehalose-6-phosphate synthase）gene,Me TPS9,was cloned from cassava. This gene had a 2598-bp open reading frame,encoded 865 amino acids,and contained a TPS conserved domain. Phylogenetic analysis revealed that Me TPS9 had close genetic relationshipwith its homologs from Capsella grandiflora and Arabidopsis thaliana,and the sequence similarity was up to 76.2% and 75.6%,respectively.Gene structural variation demonstrated that a total of 66 mutation sites were identified between wild and cultivated cassava species,of which 11 were mis-sensed. Promoter assay showed that Me TPS9 contained drought-related motif MBS,cold-related motif LTR,heat-related motif HSE,and ABA-related motif ABRE. q RT-PCR analysis revealed that Me TPS9 expressed the highest in fibrous root but the lowest in leaf. In addition,the expression of Me TPS9 was significantly induced by drought,cold,and shade stress. Me TPS9 played a regulatory role in abiotic stress suchas drought,cold,and shade treatment at the transcriptional level and it could be served as a candidate to further study its functions in abioticstress defense in cassava.