铁皮石斛DORCA基因的克隆及表达分析
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河南省科技攻关项目(092102110128);郑州市科技攻关项目(141PPTGG420)


Cloning and Expression Analysis of DORCA Gene from Dendrobium officinale
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    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1 724 bp,包含1 317 bp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。

    Abstract:

    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 DORCA closed to ZaRCA(AAK25801.1).Bioinformatics analysis showed that DORCA belonged to β isoform of RCA protein and had high identities with other plant RCAs.The DORCA contained a predicted 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 h dark and 12 h natural light photoperiods,maximal and minimal DORCA mRNA expression levels were detected at 8:30 and 20:30,respectively,which indicated that DORCA has the obvious characteristics of light-induced expression.

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崔 波,王洁琼,宋彩霞,等.铁皮石斛DORCA基因的克隆及表达分析[J].西北植物学报,2016,36(1):23-29

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  • 在线发布日期: 2016-02-24
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