辣椒高赖氨酸蛋白基因Cflr全长cDNA的克隆及其组织表达特征  

Cloning and Expression Characterization of a Lysine-rich Protein cDNA (Cflr) from Pepper

作　　者：孙晓波[1] 房瑞[1] 余桂红[1] 刘全兵[1] 马鸿翔[1] 

机构地区：[1]江苏省农业科学院农业生物技术研究所,南京210014

出　　处：《园艺学报》2008年第9期1310-1316,共7页Acta Horticulturae Sinica

基　　金：江苏省自然科学基金项目(BK2006158)

摘　　要：为了在茄科植物中寻找新的高赖氨酸蛋白基因,以辣椒栽培种‘江蔬7号’的成熟花粉为材料,根据马铃薯和番茄中高赖氨酸蛋白基因的保守序列设计引物,通过RT-PCR技术扩增到一条长390bp的cDNA片段,继而应用RACE技术克隆了一个辣椒高赖氨酸蛋白基因的全长cDNA,命名为Cflr（Gen-Bank登录号：EU367999）。Cflr基因cDNA全长920bp,5′端有84bp的非翻译区,3′端具有完整的polyA尾,包含一个编码223个氨基酸的完整开放阅读框。Cflr基因与马铃薯和番茄高赖氨酸蛋白基因cDNA序列的同源性为50%-60%,氨基酸序列的同源性为40%-50%。该基因所编码的蛋白质中赖氨酸含量为21.2%,苏氨酸含量为10.3%,是目前已知赖氨酸含量最高的天然蛋白。半定量RT-PCR结果表明,Cflr基因在辣椒未成熟花药、成熟花粉、花瓣、茎、叶片和根中均有表达,在成熟花粉和花瓣中的表达丰度最高,在叶片中表达量明显减少,而在未成熟花药、茎和根中的表达量极少。To obtain a new lysine-rich protein gene from species of solanaceae, a 390 bp fragment was amplified from pepper cultivar ＇ Jiangshu 7＇ using its matural pollen cDNA as the template and the conservative sequences of potato and tomato lysine-rich protein genes as the primers by RT-PCR. A full-length cDNA with completed open reading frame of 223 amino acids was cloned using the strategy of RACE （ rapid amplifi- cation of cDNA ends）. This cDNA was designated as Cflr （GenBank accession number： EU367999 ）, which contained 920 bp with an untranslated region of 84 bp at 5＇end and a polyA tail at the 3＇end. BLAST search against NCBI showed that the Cflr gene shared 50% -60% identity with the lysine-rich protein genes from po- tato and tomato in nucleotide and 40% - 50% in amino acid. The lysine content of CFLR protein was 21.2%, which was higher than the reported natural lysine-rich proteins, and the threonine content was 10. 3%. Analysis of semi-quantitative RT-PCR indicated that Cflr gene was transcribed in matural pollen and petal largely, less in leaf, and hardly in immature anther, stem and root.

关 键 词：辣椒 高赖氨酸蛋白基因 RACE 组织表达特征 

分 类 号：S641.3[农业科学—蔬菜学]