文心兰OnCOBRA基因克隆及表达模式分析  

Molecular Cloning and Expression Pattern of OnCOBRA Gene in Oncidium

作　　者：李玲[1] 赖钟雄[1] 陈裕坤[1] 练从龙[1] 林鹤延 

机构地区：[1]福建农林大学园艺植物生物工程研究所,福建福州350002

出　　处：《热带作物学报》2014年第8期1551-1558,共8页Chinese Journal of Tropical Crops

基　　金：福建省农业科技平台(No.2008N2001);国家科技支撑计划(No.2007BAD07B03)

摘　　要：以文心兰试管苗为材料,采用RT-PCR结合RACE法,克隆文心兰OnCOBRA基因的cDNA全长和DNA序列。结果表明:COBRA全长为1 601 bp,开放阅读框(ORF)为1 386 bp,共编码461个氨基酸;OnCOBRA的DNA序列共2 949 bp,且含有6个外显子和5个内含子。生物信息学结果表明,OnCOBRA属于不稳定的疏水蛋白,具有信号肽、跨膜结构和CCVS保守区域,亚细胞定位于细胞膜中;与无油樟、玉米、籼稻、拟南芥等具有较高的同源性。系统进化树分析结果表明,文心兰OnCOBRA蛋白与玉米(ZmCOBRA)、无油樟(AtCOBRA)、籼稻(OsCOBRA)处于统一分枝,推测OnCOBRA基因是COBRA基因家族的成员。qPCR结果表明,OnCORBA为组成型表达,在成苗期表达量最高,在文心兰类原球茎时期表达量最低。The RT-PCR combined with RACE method was used to clone the complete cDNA sequences and DNA sequences of OnCOBRA from Oncidium vitro plant. The complete eDNA sequence of OnCOBRA was 1 601 bp, encoding 461 amino acids, the DNA sequence of OnCOBRA was 2 949 bp which had 6 exons and 5 introns. The sequences of nucleotides of OnCOBRA and amino acids of OnCOBRA was high homologous with those of the known COBRA genes in other species. OnCOBRA located in plasma membrane, which had a signal peptide, transmembrane structure and CCVS protein domain. Phylogenetic tree of COBRA in plants indicated that OnCOBRA, ZmCOBRA, AtCOBRA, and OsCOBRA belonged to the same branch, and It putatively belonged to COBRA family, qPCR results indicated that OnCOBRA was constitutive expression and showed approximately a inverted ＂V＂ curve, it expressed at the highest levels in the seedings and the lowest levels in the PLB.

关 键 词：文心兰 COBRA 克隆 生物信息学 qPCR 

分 类 号：S682.31[农业科学—观赏园艺]