毛果杨HDA902基因的克隆及功能分析

发布时间：2018-02-15 14:22

  本文关键词： 毛果杨 组蛋白去乙酰化 基因克隆 载体构建 遗传转化 抗寒性　出处：《东北林业大学》2016年硕士论文　论文类型：学位论文

【摘要】：组蛋白去乙酰化酶(histone deacetylases, HDACs)催化组蛋白去乙酰化,参与组蛋白乙酰化状态的调节,对染色体结构修饰和基因表达调控发挥着重要的作用。为了研究毛果杨HDA902基因的功能,采用PCR的方法克隆了毛果杨组蛋白去乙酰化酶基因HDA902的编码序列,利用生物信息学方法分析了该基因所编码的蛋白质,并构建了GFP-HDA902融合基因表达载体,通过其在洋葱表皮细胞的表达来确定HDA902蛋白的亚细胞定位。研究结果表明：(1)毛果杨HDA902基因的开放阅读框为1500 bp,编码一个由499个氨基酸残基组成的蛋白质。(2)毛果杨HDA902蛋白与其他植物同源蛋白具有一个保守的结构域HDAC3。毛果杨HDA902蛋白在进化上与桃(Prunus persica)HDAC(XP 007200978.1)和苹果(Malus domestica)HDA19-like(XP 008348955.1)的亲缘关系较近。(3)毛果杨HDA902基因的启动子序列包含5UTR Py-rich stretch、ARE、ACE和HSE等多个与逆境和光响应相关的顺式作用元件。(4)毛果杨HDA902蛋白主要定位于细胞质中。(5)HDA902基因在毛果杨叶、茎和根三个组织中均有表达,但在茎和叶中表达量较高,根中的表达量相对较低。HDA902基因的表达受非生物胁迫的调节,盐胁迫降低了HDA902基因的表达,而低温胁迫诱导了HDA902基因的表达。(6)构建植物过量表达载体pROK Ⅱ-HDA902,通过农杆菌介导法遗传转化到烟草中,对其抗逆性功能进行分析发现,毛果杨HDA902基因在烟草中的表达提高了转基因植株的抗寒性。
[Abstract]:Histone deacetylases (HDACs) catalyze histone deacetylation, participate in the regulation of histone acetylation state, and play an important role in chromosome structure modification and gene expression regulation. The coding sequence of histone deacetylase gene (HDA902) of Populus tomentosa was cloned by PCR. The protein encoded by this gene was analyzed by bioinformatics, and the expression vector of GFP-HDA902 fusion gene was constructed. The subcellular localization of HDA902 protein was determined by its expression in onion epidermis cells. The results showed that the open reading frame of HDA902 gene of Populus tomentosa was 1500 BP, encoding a 499-amino acid residue protein. HDA902 protein has a conserved domain with other plant homologous proteins, HDAC3.The phylogenetic relationship of HDA902 protein with Prunus persica)HDAC(XP 0077978.1) and Malus domestica)HDA19-like(XP 008348955.1) is closer.; the promoter sequence of HDA902 gene of Populus tomentosa contains 5UTR Py-rich stretchard ACE and ACE. HSE and other cis-acting elements related to stress and light response. 4) the HDA902 protein of Populus tomentosa was mainly located in the cytoplasm of P. hirsutum HDA902 gene in poplar leaves. The expression of HDA902 gene in stem and root was lower than that in stem and leaf, and the expression of HDA902 gene was regulated by abiotic stress. Salt stress decreased the expression of HDA902 gene. The expression of HDA902 gene was induced by low temperature stress. PROK 鈪，

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