【作者】 刘璐；

【导师】 何光源；

【作者基本信息】 华中科技大学 ， 生物化学与分子生物学， 2007， 硕士

【摘要】 类胡萝卜素(carotenoids)是自然界存在的一类由异戊二烯为结构单元组成的化合物的一大类家族的统称。该类化合物广泛分布于部分细菌、真菌、植物等生物体内。类胡萝卜素主要分为胡萝卜素及其氧化物叶黄素两大类。很多类胡萝卜素跟人类的健康有重要的关系,特别是β-胡萝卜素以及番茄红素对眼睛的发育、抵御癌症、提高免疫力均有很重要的作用。绝大多数类胡萝卜素是C40类的碳氢四萜类(胡萝卜素)及其含氧衍生物(叶黄素),它们通过类异戊二烯途径中的部分途径合成。在高等植物中类胡萝卜素的合成途径分为两条:一条称MVA途径,另一条称MEP途径。ζ-胡萝卜素去饱和酶(ZDS)是类胡萝卜素生物合成途径上游控制β-胡萝卜素生成的关键酶,它催化9,9’-双顺-ζ-胡萝卜素脱氢生成7,9,7’,9’-四顺式-番茄红素。本研究以小麦(Triticum aestivum)叶片为材料,应用RT-PCR及RACE的方法成功克隆了小麦ZDS蛋白基因的完整阅读框序列。该开放阅读框(ORF)由1,707个碱基组成,编码568个氨基酸残基组成的蛋白质,理论分子量约为62.5 kDa,等电点为6.80,N-端有一个包含一个典型的氨基氧化酶的结构域;二级结构预测表明,不规则盘绕和α-螺旋是ZDS蛋白中主要的结构元件;同源序列比对表明其与多种植物zds基因具有较高的序列同源性,其中水稻(Oryza sativa)、玉米(Zea mays)、拟南芥(Arabidopsis thaliana)、胡萝卜(Daucus carota)同源性分别为92%、93%、83%、88%。。证实该基因确为zds基因。进化树分析显示小麦ZDS与玉米和水稻ZDS有很高的亲缘关系。该研究结果为进一步研究该基因的表达、功能和通过基因工程手段提高小麦营养价值打下了良好的基础。更多还原

【Abstract】 Carotenois are a large group of natural products that are most formally built from isoprenic unit. Those kinds of compounds are biosynthesized by some bacterium, fungi, and plants. Carotenois mainly consist of two large groups: carotenes and its oxidative derivatives(xanthophylls). Many of these carotenois are closely related to human health, especially theβ-carotene and lycopene are very important to the development of eyes, prohibiting cancers and elevating immunity. They are synthesized through part of the isoprenoids pathway. In higher plant their biosynthesis pathway includes two :one is MVA pathway,the other is MEP pathway.ζ-catotene desaturase(ZDS) is a importanr enzyme that controls the biosynthesis ofβ-carotene upstream. It catalyses the conversion of 9,9,-di-ζ-catotene to 7,9,7,9,-tetra-lycopene.In this work, reverse transcription-PCR (RT-PCR) and rapid amplification of cDNA ends (RACE) techniques had been applied to clone the total open reading frame of the zds gene using wheat (Triticum aestivum) leaves as the target material. The full-length of open reading frame is 1,707 bp and encodes a peptide containing 568 amino acid residues, with a theoretical molecular weight of 62.5 kDa, an isoelectric point of 6.80 and an amino oxidase domain at its N-terminal. Secondary structure prediction reveals thatα-helix and random coil are the main structural conformations in ZDS protein. Through sequence analysis by BlastN online, the wheat zds nucleiotide sequence showed highly homologous to the genes which are from other plant species. The homology that compared with Oryza sativa, Zea mays, Arabidopsis thaliana, Daucus carota were 92%, 93%, 83% and 88% respectively. The phylogenetic tree showed that wheat ZDS was closely related to rice ZDS and maize ZDS.The cloning and analysis of wheat zds gene has layed a good basis for research on functional expression and potential application in improving the nutrition value of wheat through genetic engeering.更多还原