RNA干扰作用对小鼠脾淋巴细胞分泌TNF-α的影响

发布时间：2018-02-27 19:18

本文关键词： RNA干扰 siRNA 转录后基因沉默 TNF-α LPS　出处：《中国医科大学》2005年硕士论文　论文类型：学位论文

【摘要】：目的肿瘤坏死因子(tumor necrosis factor-α,TNF-α)由活化的巨噬细胞和T细胞产生,分跨膜型和分泌型,是一类重要的细胞因子。TNF-α具有广泛的生物学活性,包括对造血、免疫和炎症的调节,对血管和凝血的影响以及对多种器官的作用。早期研究发现TNF-α能诱导肿瘤细胞坏死或凋亡,后来发现TNF-α是介导炎症反应、细胞免疫应答及肿瘤免疫的主要细胞因子。目前已知与TNF-α有关的疾病包括:AIDS、贫血、自身免疫性疾病、肿瘤、出血性休克、移植排斥反应、结核病、白血病、糖尿病、类风湿性关节炎等。TNF-α参与了众多疾病的发生发展过程,因此在不同水平上阻断TNF-α的作用,有可能对与TNF-α有关的疾病产生治疗作用。RNA干扰是利用具有同源互补序列的双链RNA诱发序列特异性的转录后基因沉默现象,它可以通过抑制蛋白表达模拟基因敲除技术。这一天然而古老的生物途径,是多种种属中普遍存在的抵御病毒入侵、调控基因表达的机制。抑制效果的高效性、严格的序列特异性是该技术的突出特点。从利用体外合成双链RNA到通过利用质粒稳定表达小型干扰RNA诱发RNA干扰现象,这项技术被不断完善,并被广泛的应用。在研究哺乳动物基因功能、病毒的防治、肿瘤的基因治疗等方面提供了有力的工具和新的手段。本研究从C_(57)BL/6鼠TNF-α基因序列中筛选出五个靶位点,对其分别进行RNA干扰实验。目的在于通过siRNA对TNF-α靶基因的抑制,实现对TNF-α分泌的调控。并通过分别针对五个靶位点的siRNA的干扰作用,快速筛选敏感的抑制位点,为今后克隆质粒提供最强的干扰序列。本文就位点2的干扰作用加以论述。
[Abstract]:Purpose. Tumor necrosis factor necrosis factor- 伪 (TNF- 伪) is produced by activated macrophages and T cells, transmembrane and secretory. TNF- 伪, an important cytokine, has a wide range of biological activities, including the regulation of hematopoiesis, immunity and inflammation. The effects of TNF- 伪 on blood vessels and coagulation and on various organs. Early studies have found that TNF- 伪 can induce necrosis or apoptosis of tumor cells, and later it was found that TNF- 伪 mediates inflammation. Major cytokines associated with cellular immune response and tumor immunity. Known diseases associated with TNF- 伪 include: AIDS-, anemia, autoimmune diseases, tumors, hemorrhagic shock, transplant rejection, tuberculosis, leukemia, diabetes, Rheumatoid arthritis and so on. TNF- 伪 is involved in the occurrence and development of many diseases, so at different levels of blocking the role of TNF- 伪, It is possible to treat TNF- 伪 related diseases. RNAi is a sequence-specific post-transcriptional gene silencing phenomenon induced by double-stranded RNA with homologous complementary sequences. This natural and ancient biological pathway is a common mechanism to resist virus invasion and regulate gene expression in a variety of species. Strict sequence specificity is a prominent feature of this technique. From the synthesis of double-stranded RNA in vitro to the phenomenon of RNA interference induced by stable expression of small interfering RNA by plasmid, this technique has been continuously improved. It provides powerful tools and new methods for studying mammalian gene function, virus prevention and treatment, tumor gene therapy, and so on. In this study, five targets were screened from CSP 57 / 6 mouse TNF- 伪 gene sequence. The aim of this experiment was to control the secretion of TNF- 伪 by inhibiting the target gene of TNF- 伪 by siRNA, and to quickly screen sensitive inhibitory sites by interfering with siRNA at five target sites. To provide the strongest interference sequence for cloning plasmids in the future, the interference effect of site 2 is discussed in this paper.
【学位授予单位】：中国医科大学
【学位级别】：硕士
【学位授予年份】：2005
【分类号】：R392

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