铁皮石斛RCA2基因克隆与生物信息学分析

Cloning and bioinformatics analysis of RCA2 gene in Dendrobium officinale

关键词:  铁皮石斛， RCA2基因， 克隆， 生物信息学分析

Key words:  Dendrobium officinale, RCA2 gene, gene cloning, bioinformatics analysis

摘要:

核酮糖1,5 二磷酸羧化酶/加氧酶是光合碳同化作用的关键酶，它对调节铁皮石斛Rubisco活性和光合速率具有直接的作用，对其进行基因等方面的研究，会对改变植物光合速率打下良好基础。该研究以一年生铁皮石斛叶片为材料，采用 RT PCR和 RACE技术成功克隆了Rubisco活化酶（RCA）基因，并对其进行生物信息学分析。结果表明：RCA基因全长1 730 bp，命名为RCA2(GenBank登录号KT205842)，其中5′ UTR 81 bp 、3′ UTR 326 bp，开放阅读框1 323 bp，编码440个氨基酸，分子质量为48.53 kDa，等电点为6.19，包含P loop NTPase 超家族基因结构。氨基酸多重序列比对发现RCA2核苷酸序列与蝴蝶兰的相似性高达87%。RCA2编码蛋白为亲水性蛋白；亚细胞定位于叶绿体基质；蛋白质二级结构分析，α螺旋占30.68%，延伸链占25.45%，不规则折叠占43.86%。RCA2编码蛋白质的功能预测发现，RCA2在中间代谢起到了一个非常重要的角色。该研究发现对铁皮石斛光合作用关键酶Rubisco 的分析可为其光合作用特性的发掘提供理论基础，并为提高温室大棚栽培效率提供理论依据。

Abstract:

Ribulose 1,5 bisphosphosphate carboxylase/oxygenase was a key enzyme of photosynthetic carbon assimilation, it had direct adjustive effects Rubisco activity and photosynthetic rate of in Dendrobium officinale, and laid a good foundation for changing plant photosynthetic rate by studying the gene. RCA gene was cloned from leaf of annual D. officinale using RT PCR and RACE, and was analyzed by bioinformatics. The results showed that the full length cDNA of RCA was 1 730 bp length, the cloned RCA gene was named RCA2(GenBank accession No. KT 205842). It was contained a 5′ untranslated region (5′ UTR) (81 bp), 3′ UTR (326 bp)，and an opening reading frame (ORF) (1 323 bp), which could be translated into a 410 amino acid putative peptides with a molecular weight of 48.53 kDa and a theoretical pI of 6.19, with a domain of P loop_NTPase super family. Multiple sequence alignment results showed that the homology of nucleotide sequence between the RCA2 and the reported Phalaenopsis genes was 87%. RCA2 protein was belong to hydrophilic protein，sub cellular localization was in the chloroplast. The RCA2 protein secondary structure components showed that a helix(h), extended strand(e), and random coil (c) accounted for 30.68%, 25.45% and 43.86%. RCA2 encoding protein function prediction showed that RCA2 played a very important role in intermediary metabolism. This study of the key enzyme photosynthesis Rubisco in Dendrobium officinale provides the information for the research of the photosynthetic characteristics, and the basis for improving the efficiency of greenhouse cultivation.