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中国卤虫滞育胚胎解除过程中hyou1基因的分子克

发布时间:2018-06-29 12:49

  本文选题:中国卤虫 + hyou1 ; 参考:《辽宁师范大学》2016年硕士论文


【摘要】:中国卤虫(Artemia sinica)广泛分布于高盐水体中,遇到环境胁迫可以产生休眠卵,其胚胎滞育期新陈代谢减慢,包囊可以抵御低氧、高盐、寒冷等恶劣环境的影响。缺氧上调蛋白Hypoxia up-regulated 1,也被称为Orp150,其属于热休克蛋白家族的成员之一。Hypoxia up-regulated 1在低氧条件下的大鼠胶质细胞中首次被发现,积累在缺氧条件下的内质网内,被认为在蛋白质折叠和分泌中发挥重要作用。本研究采用RACE技术首次得到了中国卤虫Hypoxia up-regulated 1基因的cDNA全长As-hyou1,同时也对As-hyou1进行了生物信息学分析和分子进化等方面的分析。As-hyou1的基因全长为2952bp,As-hyou1基因包含了116bp的5’非编码区以及180bp的3’非编码区,其中2656bp为开放阅读框结构。As-hyou1基因所编码的HYOU1蛋白是一含有886个氨基酸残基的蛋白,具有典型的PTZ00009结构域,属于NBD糖激酶HSP70-actin超家族成员。理论分子量100.5KD,等电点为5.34。Signal P4.1检测到蛋白序列中含信号肽序列,故中国卤虫HYOU1为分泌性蛋白。利用原位杂交技术对As-hyou1的表达部位进行了分析,As-hyou1广泛分布于胚胎发育的各时期。卤虫虫体生长发育至已分化阶段,卤虫的头胸部、腹部、体表、附肢都会检测到阳性信号。采用荧光实时定量PCR,对As-hyou1的表达模式进行了研究,结果显示,As-hyou1在卤虫胚胎滞育刚刚结束阶段也就是滞育卵重新回到其正常发育进程时具有较高的表达(0h),在早期胚胎发育期逐渐下调(5h-15h),在虫体发育到无节幼体时期表现出上升趋势;在高盐刺激下As-hyou1的表达由50‰开始逐步下调;同时低温刺激下表达量呈下降趋势。研究初步证明,As-hyou1介导调节抑制由缺氧引起的细胞程序性死亡,并通过内质网促进翻译、转化和处理其他相关蛋白。本研究对进一步阐明As-hyou1在中国卤虫滞育解除、早期胚胎发育所起的作用,及对中国卤虫滞育发生与解除现象的分子机制的研究具有较为重要的理论意义。
[Abstract]:The Artemia Sinica in China is widely distributed in high salt water bodies. It can produce dormant eggs during environmental stress. The metabolism of the embryo diapause period is slow and the capsule can resist the adverse environment of hypoxia, high salt and cold. The anoxic up protein Hypoxia up-regulated 1, also known as Orp150, is a member of the heat shock protein family. .Hypoxia up-regulated 1 was first found in rat glial cells under hypoxic conditions, accumulated in the endoplasmic reticulum under anoxic conditions, and considered to play an important role in protein folding and secretion. In this study, the total cDNA length As-hyou1 of the Hypoxia up-regulated 1 gene of Chinese halus was first obtained by RACE technology, and it was also used in the study. As-hyou1 carried out the analysis of bioinformatics and molecular evolution. The whole length of.As-hyou1 was 2952bp, and the As-hyou1 gene contained the 5 'non coding region of 116bp and the 3' non coding region of 180bp, and the HYOU1 protein encoded by 2656bp for the open reading frame structure.As-hyou1 was a protein containing 886 amino acid residues. It has a typical PTZ00009 domain, belonging to the NBD glycokinase HSP70-actin superfamily. The theoretical molecular weight is 100.5KD, and the isoelectric point is 5.34.Signal P4.1 to detect the sequence of signal peptide in the protein sequence, so the Chinese Artemia HYOU1 is a secretory protein. In situ hybridization technique is used to analyze the expression of As-hyou1, and As-hyou1 is widely distributed. At all stages of embryonic development. The body of the Artemia grew to the stage of differentiation, and the positive signals were detected in the head, abdomen, body surface and appendage of the halus. The expression pattern of As-hyou1 was studied by real time fluorescence quantitative PCR. The results showed that the diapause embryo of the haloembryo embryo was just returned to the diapause egg. In the normal development process, it has a higher expression (0h), decreasing in the early embryonic development period (5h-15h), and showing an upward trend in the period of the development of the insect body to the nauplite. Under the high salt stimulation, the expression of As-hyou1 is gradually downregulated from 50 per thousand. At the same time, the amount of the surface reaches a downward trend at the same time. The study has preliminarily proved that As-hyou1 mediated modulation. It inhibits the programmed cell death caused by hypoxia and promotes translation, transformation and processing of other related proteins through endoplasmic reticulum. This study is more important for further clarifying the role of As-hyou1 in the diapause and early embryonic development of the Chinese halus, as well as the molecular mechanism of the diapause and disintegration of the halus in China. The theoretical significance.
【学位授予单位】:辽宁师范大学
【学位级别】:硕士
【学位授予年份】:2016
【分类号】:Q78;Q954.4

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