香鳞毛蕨1-脱氧-D-木酮糖-5-磷酸合酶基因的克隆及表达分析

Cloning and Expression Analysis of 1-Deoxy-D-Xylitose-5-Phosphate Dynthase Gene from Dryopteris fragrans

1-脱氧-D-木酮糖-5-磷酸合酶(DXS)是甲基-D-赤藓醇-4-磷酸(MEP)途径中控制影响植物萜类化合物合成的第一个限速酶。该研究对香鳞毛蕨(Dryopteris fragrans)DfDXS基因进行序列特征及生物信息学分析,并通过qRT-PCR技术分析其在外源激素、干旱、盐胁迫、高温及低温处理下的表达模式,旨在探究DfDXS基因在香鳞毛蕨萜类生物合成及抗逆机制中的作用,为进一步解析香鳞毛蕨抗逆分子机制奠定基础。结果显示:(1)DfDXS1基因全长2139 bp,编码712个氨基酸,而DfDXS2全长2160 bp,编码719个氨基酸;结构域分析显示,其具有典型的转酮醇酶保守域,包含焦磷酸硫胺素结合位点和转酮醇酶结构域;DfDXS氨基酸序列与江南卷柏(Selaginella moellendorffii)和银杏(Ginkgo biloba)的DXS等关系较近。(2)水杨酸(SA)处理下,DfDXS基因的相对表达量先升高后降低;脱落酸(ABA)抑制DfDXS的表达;DfDXS1/2在茉莉酸甲酯(MeJA)处理下相对表达水平均显著高于对照;乙烯利(Eth)抑制DfDXS的表达,但DfDXS1处理3 h时表达水平显著高于对照。(3)聚乙二醇(PEG)、高温和低温均诱导DfDXS1上调表达。研究推测,香鳞毛蕨DXS基因在萜类物质合成与逆境胁迫机制中发挥着重要的作用。

1-deoxy-D-xylose-5-phosphate synthase(DXS)is the first rate-limiting enzyme that controls the synthesis of plant terpenes in the methyl-D-erythritol-4-phosphate(MEP)pathway.In this study,the sequence characteristics and bioinformatics of DfDXS gene of Dryopteris fragrans were analyzed,and its expression patterns under exogenous hormone,drought,salt stress,high and low temperature treatments were analyzed by qRT-PCR technique,for exploring the role of DfDXS gene in terpenoid biosynthesis and stress resistance mechanism.The results showed that:(1)DfDXS1 full-length 2139bp encodes 712 amino acids,and DfDXS2 full-length 2160 bp encodes 719 amino acids.Domain analysis showed that it had a typical conservative domain of transketolase,including thiamine pyrophosphate binding site and transketolase domain.The genetic relationship is close to that of Selaginella moellendorffii DXS,Ginkgo biloba DXS1.(2)Under the treatment of salicylic acid(SA),the relative expression of DfDXS increased at first and then decreased;abscisic acid(ABA)inhibited the expression of DfDXS;DfDXS1 and DfDXS2 re-sponded strongly to methyl jasmonate(MeJA)treatment,and the relative expression level of DfDXS1 was significantly higher than that of the control;Ethephon(Eth)inhibited the expression of DfDXS,but the expression of DfDXS1 was higher than that of the control at 3 h.(3)Polyethylene glycol(PEG),high temperature and low temperature all up-regulated the expression of DfDXS1.The results showed that DXS gene plays a role in terpenoid synthesis and stress mechanism.This study provides a basis for further analysis of the molecular mechanism of stress resistance of D.fragrans.