铁皮石斛尿苷二磷酸葡萄糖焦磷酸化酶基因的克隆与分析

Cloning and Expression Analysis of UGPase Gene in Dendrobium Officinale Kimura et Migo

目的:研究尿苷二磷酸葡萄糖焦磷酸化酶(UGPase)在铁皮石斛不同组织中的表达情况,探讨其与石斛多糖累积的关系。方法:根据铁皮石斛原球茎转录组数据中的DoUGPase1序列设计引物,并利用RACE技术克隆该基因;用生信分析软件预测其蛋白结构,并进行系统发育分析;利用实时荧光定量PCR(q RT-PCR)技术研究该基因在铁皮石斛不同年份,不同组织中的表达。结果:通过RACE技术得到1个3054 bp基因,命名为DoUGPase1,其ORF长1530 bp,编码510个氨基酸残基,与油棕(Elaeis guineensis)和波斯枣(Phoenix dactylifera)有一定的同源性,且DoUGPase1在3年生的植株中相对表达量整体较高(P<0.05)。结论:我们预测在铁皮石斛的生长发育过程中DoUGPase1基因与多糖累积的活跃程度成正比例。

Objective： To observe the expression of UGPase in Dendrobium, and investigate its relationship with Dendrobium polysaccharide accumulation. Methods： The primers were designed according to the transcriptomic UGPasel tmigene sequence ofproto- corm in D. officinale, then DoUGPase 1 transcription factors were cloned by RACE techniques. Through bioinformatics analysis and phy- logenesis analysis, the protein structure was predicted, and its affinity was obstained. The relative expression of D.officinale in different years and different tissues was analyzed through Roche Light circle 96 real-time fluorescent quantitative PCR （RT-qPCR） experiment. Results： By RACE technique, a new 1530 bp long gene named DoUGPasel was obstained. Its ORF （Open reading frame） was 1530 bp encoding 510 amino acids. The DoUGPasel had the relatives with Elaeis guineensis and Phoenix dactylifera. The DoUGPasel expres- sion in 3 years plant was the biggest（P〈0.05 ）. Conclusions： The expression level of DoUGPaselgene might be a positive proportion with active degree of dendrobium polysaccharide accumulation in the growth and development of dendrobium officinale.