月季真核生物起始因子5aei中f5a基因的功四能研究.docx

摘 摘 要 真核起始因子5A(elF5A)是目前发现的唯一一种含有特殊氨基酸 hypusine(g}t咯赖氨酸)的高度保守性蛋白因子。已有的研究表明其参与翻译的延 伸、mRNA的转运及降解、细胞的生长发育以及程序性死亡等多种代谢途径。 本研究通过对月季(Rosa Chinensis)elF5A基因(尺ce伊51。4)进行洋葱表皮亚细 胞定位，结果显示RcelF5A主要定位于细胞质中。用RT-PCR方法研究该基因在 多种非生物胁迫下的表达模式，初步推断该基因与高温、氧化和渗透胁迫有关。 于是将该基因转入真核模式生物毕氏酵母(Pichia pastoris)SMDll68中，通过 各种胁迫处理及生长势测定分析证明该基因的表达能够提高宿主菌的高温和氧 化胁迫抗性。进一步将该基因转入模式植物拟南芥中，通过对不同条件下植株的 表型及生理指标分析，证明过表达该基因能够提高阳性植株的高温、氧化及渗透 胁迫的抗性。 关键词：elF5A，月季，毕氏酵母，拟南芥，非生物胁迫抗性 中图分类号：Q 78 AbstractEukaryotic Abstract Eukaryotic translation initiation factor 5A(elF5A)is the only cellular protein known to contain the unusual amino acid hypusine．It is a highly conserved protein found in all eukaryotic organisms．Although originally identified as a translation initiation factor，recent studies suggest that elF5A is mainly involved in translation elongation，mRNA turnover and decay，cell proliferation，and programmed cell death． However，the precise cellular function of elF5A remains largely unknown，especially in plants．In this work，we characterized the RcelF5A from Rosa chinensis． Subcellular localization of RcelF5A showed that it localized predominantly to the cytoplasm of onion epidermal cells．RcelF5A expression is up-regulated in Rosa chinensis under high temperature，oxidative and osmotic stress conditions． Recombinant RcelF5A was overexpressed in Pichia pastoris strain SMDl 168 to study its possible function under stress conditions．The recombinant SMDl 168 cells that accumulated RcelF5A showed improved viability under thermal and oxidative stress conditions compared to control cultures．We then produced transgenic Arabidopsis that constitutively enhanced or suppressed expression of RcelF5A．The RcelF5A over-expression plants exhibited increased resistance to heat，oxidative and osmotic stresses，while the suppressed expression plant(three AtelF5A isoforms in Arabidopsis were down-regulated)showed more susceptibility to these stresses． Keywords：elF5A，Rosa chinensis，Pichia pastori，Arabidopsis thaliana，abiotic stress resi