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Plant Diversity ›› 2015, Vol. 37 ›› Issue (01): 71-77.DOI: 10.7677/ynzwyj201514057

• 研究论文 • 上一篇    下一篇

樟叶越桔熊果苷合成酶基因VdAS1的克隆及序列分析

 杜维1, 丁勇1**, 朱东阳2, 尹继庭2, 刘小烛1, 赵平2   

  1. 1 西南林业大学生命科学学院,昆明650224;2 西南林业大学西南山地森林资源保育
    与利用省部共建教育部重点实验室,昆明650224
  • 收稿日期:2014-04-03 出版日期:2015-01-25 发布日期:2014-06-13
  • 基金资助:

    国家自然科学基金地区科学基金项目 (21462040, 31460076);云南省优势特色重点学科生物学一级学科建设项目 (50097505)

Cloning and Bioinformatics Analysis of VdAS1 Gene in Vaccinium dunalianum (Ericaceae)

 DU  Wei-1, DING  Yong-1**, ZHU  Dong-Yang-2, YIN  Ji-Ting-2, LIU  Xiao-Zhu-1, ZHAO  Ping-2   

  1. 1 College of Life Sciences, Southwest Forestry University, Kunming 650224, China; 2 Key Laboratory for Forest
    Resources Conservation and Use in the Southwest Mountains of China, Ministry of Education,
    Southwest Forestry University, Kunming 650224, China
  • Received:2014-04-03 Online:2015-01-25 Published:2014-06-13
  • Supported by:

    国家自然科学基金地区科学基金项目 (21462040, 31460076);云南省优势特色重点学科生物学一级学科建设项目 (50097505)

摘要:

根据樟叶越桔 (Vaccinium dunalianum) 叶芽转录组测序获得的熊果苷合成酶基因VdAS1部分EST序列设计引物,结合RACEPCR技术获得VdAS1基因全长1577bp cDNA序列。其完整的开放阅读框推测编码由475个氨基酸残基组成的VdAS1蛋白,理论相对分子量为5222kD,等电点pI=574,负电荷残基 (Asp+Glu) 总数为53个,正电荷残基 (Arg+Lys) 总数为45个,不稳定系数为3793,属稳定性蛋白质。生物信息学分析表明VdAS1与枸杞的糖基转移酶相似率高达74%,其二级结构主要构件为α螺旋和随机卷曲,不存在跨膜区,属于亲水性蛋白质,并结合信号肽预测结果推测VdAS1直接锚定在细胞基质中行使功能。该研究为后期VdAS1的异源表达和功能研究奠定了基础。

关键词: 樟叶越桔, 熊果苷合成酶, 基因克隆, 生物信息学分析

Abstract:

Arbutin was widely used in cosmetics as a whitening agent because of blocking the synthesis of melanin as the inhibitor of tyrosinase. Arbutin synthase (AS) as one member of UDP glycosyltransferases (UGTs) which belongs to the glycosyltransferase (GTs) family is a ratelimiting enzyme in arbutin synthesis from hydroquinone. To reveal the biological function and provide scientific basis of application on plant genetic engineering for Vaccinium dunalianum arbutin synthase (VdAS), a gene VdAS1 encoding VdAS1 was isolated by RTPCR combined with RACEPCR from Vdunalianum. The fulllength cDNA clone comprised 1577 nucleotides consisting of a 17nucleotide 5′untranslated region, an open reading frame of 1428 nucleotides, and a 132nucleotide 3′untranslated region. The open reading frame encoded a putative VdAS1 comprising 475 amino acid residues with molecular weight of 5222kD. and isoelectric point of 574. The total numbers of negative charged residues (Asp+Glu) and positive charged residues (Arg+Lys) in VdAS1 were 53 and 45, respectively. VdAS1 shares 74% similarity with UGT in wolfberry (Lycium barbarum). It was predictive VdAS1 had no signal peptide and transmembrane structure. The main parts of predicted secondary structures of VdAS1 were ahelices and random coils. The study results established the foundation for the heterologous expression and functional analysis of VdAS1.

Key words: Vaccinium dunalianum, Arbutin synthase, Gene cloning, Bioinformatics analysis

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