小麦果聚糖合成酶基因6-SFT克隆和功能验证  

Isolation and Functional Determination of Fructan Biosynthesis Enzyme Encoding Gene 6-SFT from Common Wheat (Triticum aestivum L.)

作　　者：高翔[1,2,3] 佘茂云[1,2,3] 殷桂香 于洋[4] 别晓敏[1,2,3] 杜丽璞[1,2,3] 徐惠君[1,2,3] 叶兴国[1,2,3] 

机构地区：[1]中国农业科学院作物科学研究所 [2]国家农作物基因资源与基因改良重大科学工程 [3]农业部作物遗传育种重点开放实验室,北京100081 [4]中国农业科学院生物技术研究所,北京100081

出　　处：《科技导报》2009年第23期70-75,共6页Science & Technology Review

基　　金：中国高技术研究发展计划(863计划)项目(2007AA10Z129)

摘　　要：许多研究表明植物中果聚糖累积有助于提高植物耐逆境胁迫能力,但对小麦中果聚糖合成相关基因及其与抗逆性关系还缺乏研究。本研究拟从普通六倍体小麦中克隆出果聚糖合成酶基因6-SFT并进行生物学特性及功能研究,以期解析小麦通过累积果聚糖途径提高自身抗逆境胁迫能力的分子机理,为转6-SFT基因提高小麦抗逆境胁迫能力提供理论依据。利用touch-down PCR技术从小麦品种杨麦6(Triticum aestivum cv. Yangmai 6)基因组及cDNA中分离出编码6-SFT基因的全长序列,运用生物学信息技术对其编码蛋白的相关理化及生物学功能进行了预测,同时利用农杆菌介导法将所克隆的小麦6-SFT基因转入了烟草,对转基因烟草植株进行了抗旱、抗盐和抗低温鉴定。结果表明,小麦中6-SFT基因的gDNA和cDNA长度分别为3134bp和1851bp,序列结构分析表明该基因含有4个外显子、3个内含子,编码产物为约68kD的蛋白质,其等电点(pI)约为5.25,具有液泡定位信号;转小麦6-SFT基因烟草植株表现出较强的抗旱、抗盐和抗低温能力。本研究为利用基因工程技术改良小麦抗逆性提供重要理论依据。Previous studies have shown the relevance of fructans deposit to plant tolerance to abiotic stresses. In this study, 6-SFT, a fructan biosynthesis enzyme encoding gene, was isolated from hexaploid wheat to reveal the resistance mechanism against adverse environmental factors and to provide a theoretical basis for future transgenic researches in wheat. Touch-down PCR was adopted to isolate the gene coding for fructan biosynthesis enzyme, sucrose： fructan 6-fructosyhransferase （6-SFT）, from common wheat cv. Yangmai 6 genome DNA （gDNA） and complementary DNA （cDNA）. Relevant bioinformatics assay on 6-SFT gene was performed online and its ability to resist abiotic stresses was confirmed in transgenic tobacco plants obtained by using Agrobacterium -mediated transformation approach. The results show that the gDNA and cDNA of 6-SFT are 3 134 bp and 1 851 bp, respectively, containing four exons and three introns and encoding a protein with a MW of about 68 kD. The isoelectric point （pI） is 5.25 with potential vacuoledirected signal peptide （TLPI）. Positive transgenic tobacco lines are confirmed through traditional PCR detection and ELISA assay, showing high tolerance to drought, salinity and low temperaturestress. This study provides a theoretical platform for the improvement of tolerance to adverse environmental stresses through subsequent transgenic pathway in wheat.

关 键 词：小麦 6-SFT基因 烟草 转基因 抗逆性 

分 类 号：S512.1[农业科学—作物学]