| 摘要: |
| 【目的】克隆珍稀濒危兰科药用植物铁皮石斛（Dendrobium officinale Kimura et Migo）鲨烯合酶（Squalene synthase,SS）基因（DoSS），并对其进行生物信息学与组织表达模式分析，为研究其生物学功能提供参考。【方法】采用RT-PCR和RACE技术，获得DoSS基因全长序列；利用生物信息学软件预测DoSS蛋白的理化性质、结构域及亚细胞定位等分子特性；用DNASTAR 6.0和MEGA 4.0分别进行DoSS蛋白氨基酸多序列比对和进化关系分析；借助qPCR检测DoSS基因的组织表达模式。【结果】成功克隆获得铁皮石斛鲨烯合酶基因，命名为DoSS（GenBank注册号JX272631），其cDNA全长1 735 bp，编码一条由410个氨基酸组成的多肽，分子质量46.99 ku，等电点7.13；DoSS蛋白含有鲨烯/八氢番茄红素合成酶保守结构域（44-315位氨基酸）；DoSS与其他多种植物SS基因的编码蛋白同源性很高（61%～75%），与水稻、玉米等单子叶植物的亲缘关系较近；DoSS基因为组成型表达，在根中的表达量最大，为叶的12.41倍；茎次之，为叶的1.87倍。【结论】成功克隆得到1个鲨烯合酶基因（DoSS）全长cDNA；DoSS基因的（在根中高）表达特征暗示，其可能在铁皮石斛根的生长发育中发挥重要的调控功能。 |
| 关键词: 铁皮石斛 鲨烯合酶 基因克隆 表达模式 结构域 萜类 |
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| 基金项目:教育部科学技术研究重点项目（211177）；陕西省科技计划青年科技新星项目(2012KJXX-44)；陕西省自然科学基础研究计划项目（2011JQ4016）；陕西省教育厅专项科研计划项目（2013JK0829） |
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| Characterization of a squalene synthase gene in Dendrobium officinale Kimura et Migo |
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| Abstract: |
| 【Objective】This study was carried out to clone a squalene synthase gene (DoSS) from a medicinally important endangered orchid species Dendrobium officinale Kimura et Migo,followed by bioinformatics and expression analysis.【Method】RT-PCR and RACE approaches were used to isolate the full-length gene.Characteristics of the molecular weight,conserved domain and subcellular localization of the deduced DoSS protein were determined using a series of bioinformatics tools.The analyses of multiple alignment and phylogenetic tree were performed using DNASTAR 6.0 and MEGA 4.0,respectively.qPCR was employed to examine the tissue specific expression patterns of DoSS.【Result】A full-length cDNA encoding squalene synthase,designated as DoSS (GenBank accession No.JX272631),was identified from Dendrobium officinale Kimura et Migo.The gene comprised 1 735 bp encoding a 410 amino acid polypeptide with a calculated molecular weight of 46.99 ku and an isoelectric point (pI) of 7.13.The deduced DoSS protein contained the squalene/phytoene synthase conserved domain (44-315).DoSS was highly homologous (61%-75%) to a number of SS proteins from various plants,and was closely related to rice and maize monocots.DoSS constitutively expressed among the three studied tissues and the transcripts were the abundant in roots which were 12.41 times than in leaves, followed by stems (1.87 times than in leaves).【Conclusion】The full-length cDNA of DoSS gene was successfully cloned.The high expression level of DoSS in Dendrobium officinale Kimura et Migo roots suggested that the gene might play a vital regulatory role in the roots. |
| Key words: Dendrobium officinale Kimura et Migo squalene synthase gene cloning expression pattern domain terpenoid |
