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基于RNA-Seq技术的连翘转录组组装与分析及SSR分子标记的开发

DOI: 10.1360/N052014-00273, PP. 301-310

Keywords: RNA-Seq,转录组,次生代谢,苯乙醇苷,木脂素,SSR分子标记,连翘

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Abstract:

连翘既是传统的中药材,又是优良的城市绿化树种,具有重要的经济价值和生态价值.但是连翘基因资源非常匮乏,限制了其分子生物学和基因功能的研究.本研究以连翘根、茎、叶、花和果实等器官的混合样品作为材料,利用IlluminaHiSeqTM2500测序平台对其进行转录组测序.共获得23164327条干净数据(cleanreads),总碱基数为4678791021bp.Cleanreads经denovo组装后获得45112条unigenes.进一步利用五大公共数据库进行同源比对,注释了28699条unigenes.其中,473个基因参与了连翘次生物质的合成和代谢,包括81个与苯丙氨酸和苯丙烷代谢相关的基因.对这81个基因的分析表明,有4个基因编码苯丙氨酸脱氨酶,1个基因编码肉桂酸4-羟化酶,2个基因编码4-香豆酰:辅酶A连接酶.这3个酶催化了连翘中主要药用活性物质苯乙醇苷和木脂素前体肉桂酸衍生物的生物合成.此外,还发现了2个松脂醇-落叶松树脂醇还原酶和1个开环异落叶松脂醇脱氢酶编码基因,这2个酶是木脂素合成的关键酶.最后,分析了长度在1kb以上的12721个unigenes的基因结构,检测到3199个SSR位点,并对其中40个位点进行了验证.本研究不仅为连翘基因克隆和分子生物学研究提供了丰富的基础数据信息,而且为连翘遗传多样性研究、指纹图谱构建和分子标记辅助选育奠定了分子基础.

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