丹参多酚氧化酶基因的克隆及表达分析
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国家自然科学基金(81373908);


Cloning of Polyphenol Oxidase Gene from Salvia miltiorrhiza Hairy Roots and Its Expression Analysis
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    摘要:

    前期研究发现多酚氧化酶(PPO)能正向调控丹酚酸B合成,该研究运用RACE技术,从丹参毛状根中克隆到多酚氧化酶基因(SmPPO,GenBank登录号为KF712274)全长序列,其cDNA全长1 930 bp,开放阅读框为1 770 bp,编码589个氨基酸。将SmPPO与管状花目其它4个物种进行氨基酸序列比对,在N端类囊体转移结构域中发现都存在2个N豆蔻酰化位点。在丹参毛状根培养液中加入不同诱导因子,利用实时荧光定量PCR检测,发现该基因在酵母提取物处理中表达量显著上调,但在银离子、抗坏血酸和L半胱氨酸处理中表达受到明显抑制。运用HPLC技术同步检测毛状根中丹酚酸B含量,显示出与基因表达相同的变化趋势。研究表明,丹参中多酚氧化酶基因(SmPPO)对丹酚酸B的合成具有正向调控作用。

    Abstract:

    The polyphenol oxidase (PPO) was found to positively regulate the accumulation of salvianolic acid B (SAB). This investigation further clone its cDNA sequence from hairy roots of Salvia miltiorrhiza by using RACE technique (named as SmPPO, GenBank accession No. KF712274). The full length cDNA of SmPPO was 1 930 bp with a 1 770 bp open reading frame, encoding a protein of 589 amino acids. Two Nglycosylation sites were found in Nterminus thylakoid transfer domain, when comparing the amino acid sequences of SmPPO to other PPOs in Tubiflorae. Expression analysis of realtime PCR indicated that PPO transcript levels significantly increased in yeast extract (YE)elicited hairy roots, whereas it significantly decreased in the Ag+, ascorbic acid and cysteine treatments. Correspondingly, the accumulation of SAB was significantly increased in YEtreatment hairy roots, and was inhibited by the three elicitor treatments. These results suggest that SmPPO gene positively regulates the accumulation of SAB.

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时王珂,孙奕玥,舒志明,等.丹参多酚氧化酶基因的克隆及表达分析[J].西北植物学报,2016,36(9):1735-1742

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  • 在线发布日期: 2016-09-30
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