黄瓜贝壳杉烯氧化酶基因CKO的克隆及其表达分析  

Cloning and Expression Analysis of ent-kaurene Oxidase Gene CKO in Cucumber

作　　者：胡宏敏[1] 蒋芳玲[1] 曹雪[1] 吴震[1] 王广龙[1] 

机构地区：[1]南京农业大学园艺学院,农业部华东地区园艺作物生物学与种质创制重点实验室,南京210095

出　　处：《园艺学报》2012年第6期1131-1140,共10页Acta Horticulturae Sinica

基　　金：江苏省自然科学基金项目(BK2008342);江苏省科技支撑计划项目(BE2009403;BE2009413);苏北科技发展计划项目(BN2010054);江苏高校优势学科建设工程项目

摘　　要：以‘戴多星’黄瓜为试材,应用同源克隆和RACE技术从茎尖中得到赤霉素合成的关键酶——贝壳杉烯氧化酶(KO)基因cDNA全长序列,命名为CKO,GenBank登录号为JN792591,其长度为1 895 bp,开放阅读框(ORF)编码519个氨基酸,相对分子量为59.211 kD,等电点(PI)8.45。氨基酸同源性分析发现,CKO与苦瓜的KO同源性最高,达84%;序列结构分析表明,CKO属于细胞色素超家族P450系,具有细胞色素P450的血红素结构域FXXGXRXCXG和跨膜结构域。实时荧光定量分析表明,CKO在遮荫的徒长苗中表达量最高,是正常苗的5.18倍,在遮阴条件下喷施多效唑可降低期表达。With homologous cloning and RACE technology, ent-Kaurene oxidase （KO）, a key enzyme in the pathway of gibberellins biosynthesis, was cloned from the shoot tip of＇ Deltastal＇ （ Cucumis sativus L. ） and named CKO （GenBank accession number： JN792591 ） . The full eDNA was 1 895 bp in length with an open reading frame （ORF） encoding a protein of 519 amino acids. The protein molecular weight and isoelectric point were predicted to be 59.211 kD and 8.45 respectively. Amino acids homology analysis indicated that the sequence had 84% similarity with that of Momordica charantia. Sequence analysis showed that CKO belonged to cytochrome P450 superfamily and contained cysteine hemeiron ligand signature （FXXGXRXCXG） and transmembrane region. The fluorescent quantitative RT-PCR result showed that the expression level of CKO was the highest in leggy seedling under shading treatment, being 5.18 times of the normal one. While the expression of CKO significantly reduced after spraying paclobutrazol at shading.

关 键 词：黄瓜 贝壳杉烯氧化酶 基因 克隆 表达分析 

分 类 号：S642.2[农业科学—蔬菜学]