铁皮石斛DORCA基因的克隆及表达分析  

Cloning and Expression Analysis of DORCA Gene from Dendrobium officinale

作　　者：崔波[1,2] 王洁琼[2] 宋彩霞[2] 袁秀云[1] 蒋素华[1] 梁芳[1] 刘佳[2] 叶永忠[2] 

机构地区：[1]郑州师范学院,郑州450044 [2]河南农业大学生命科学学院,郑州450002

出　　处：《西北植物学报》2016年第1期23-29,共7页Acta Botanica Boreali-Occidentalia Sinica

基　　金：河南省科技攻关项目(092102110128);郑州市科技攻关项目(141PPTGG420)

摘　　要：为了研究铁皮石斛的光合特性,根据Rubisco活化酶(RCA)基因的保守序列设计兼并引物,采用RT-PCR和RACE方法从铁皮石斛叶中分离到一个RCA基因,命名为DORCA(GenBank登录号KT205841)。DORCA基因cDNA全长为1 724bp,包含1 317bp开放阅读框(ORF),编码438个氨基酸。系统进化分析显示,DORCA与马蹄莲ZaRCA(AAK25801.1)亲缘关系最近。生物信息学分析表明,DORCA与其他植物的RCA蛋白具有较高一致性,属于RCA的β亚基。DORCA蛋白具有定位于叶绿体的N端转运肽,2个保守的ATP结合结构域和多个磷酸化位点。实时荧光定量PCR(qRT-PCR)分析表明,DORCA基因在茎、叶中表达;在自然光周期条件下,DORCA基因在8:30时表达量最高,20:30时表达量最低,具有明显的光诱导表达特性。To study the photosynthesis of Dendrobium officinale,we isolated a full-length cDNA of Rubisco activase DORCA（Accession No.KT205841）from this plant by the method of RT-PCR and RACE using degenerate primers designed according to the conserved region of RCA.The full-length cDNA of DORCA was 1 724 bp,with an ORF of 1 317 bp encoding 438 putative amino acids.Phylogenetic analysis showed that DORCAclosed to ZaRCA（AAK25801.1）.Bioinformatics analysis showed that DORCA belonged toβisoform of RCA protein and had high identities with other plant RCAs.The DORCA contained apredicted N-terminal transit peptide to chloroplast,two high conserved ATP-binding domains and multiple phosphorylation sites.The real-time quantitative PCR（qRT-PCR）results showed that DORCA was only detected in green tissues.In the 12 hdark and 12 hnatural light photoperiods,maximal and minimal DORCA mRNA expression levels were detected at 8：30and 20：30,respectively,which indicated that DORCAhas the obvious characteristics of light-induced expression.

关 键 词：铁皮石斛 RUBISCO活化酶 分子特征 表达特性 

分 类 号：Q785[生物学—分子生物学] Q786