本文關(guān)鍵詞： RNA干擾 siRNA 轉(zhuǎn)錄后基因沉默 TNF-α LPS　出處：《中國醫(yī)科大學(xué)》2005年碩士論文　論文類型：學(xué)位論文

【摘要】：目的 腫瘤壞死因子(tumor necrosis factor-α,TNF-α)由活化的巨噬細(xì)胞和T細(xì)胞產(chǎn)生,分跨膜型和分泌型,是一類重要的細(xì)胞因子。TNF-α具有廣泛的生物學(xué)活性,包括對(duì)造血、免疫和炎癥的調(diào)節(jié),對(duì)血管和凝血的影響以及對(duì)多種器官的作用。早期研究發(fā)現(xiàn)TNF-α能誘導(dǎo)腫瘤細(xì)胞壞死或凋亡,后來發(fā)現(xiàn)TNF-α是介導(dǎo)炎癥反應(yīng)、細(xì)胞免疫應(yīng)答及腫瘤免疫的主要細(xì)胞因子。目前已知與TNF-α有關(guān)的疾病包括:AIDS、貧血、自身免疫性疾病、腫瘤、出血性休克、移植排斥反應(yīng)、結(jié)核病、白血病、糖尿病、類風(fēng)濕性關(guān)節(jié)炎等。TNF-α參與了眾多疾病的發(fā)生發(fā)展過程,因此在不同水平上阻斷TNF-α的作用,有可能對(duì)與TNF-α有關(guān)的疾病產(chǎn)生治療作用。RNA干擾是利用具有同源互補(bǔ)序列的雙鏈RNA誘發(fā)序列特異性的轉(zhuǎn)錄后基因沉默現(xiàn)象,它可以通過抑制蛋白表達(dá)模擬基因敲除技術(shù)。這一天然而古老的生物途徑,是多種種屬中普遍存在的抵御病毒入侵、調(diào)控基因表達(dá)的機(jī)制。抑制效果的高效性、嚴(yán)格的序列特異性是該技術(shù)的突出特點(diǎn)。從利用體外合成雙鏈RNA到通過利用質(zhì)粒穩(wěn)定表達(dá)小型干擾RNA誘發(fā)RNA干擾現(xiàn)象,這項(xiàng)技術(shù)被不斷完善,并被廣泛的應(yīng)用。在研究哺乳動(dòng)物基因功能、病毒的防治、腫瘤的基因治療等方面提供了有力的工具和新的手段。本研究從C_(57)BL/6鼠TNF-α基因序列中篩選出五個(gè)靶位點(diǎn),對(duì)其分別進(jìn)行RNA干擾實(shí)驗(yàn)。目的在于通過siRNA對(duì)TNF-α靶基因的抑制,實(shí)現(xiàn)對(duì)TNF-α分泌的調(diào)控。并通過分別針對(duì)五個(gè)靶位點(diǎn)的siRNA的干擾作用,快速篩選敏感的抑制位點(diǎn),為今后克隆質(zhì)粒提供最強(qiáng)的干擾序列。本文就位點(diǎn)2的干擾作用加以論述。
[Abstract]:Purpose. Tumor necrosis factor necrosis factor- 偽 (TNF- 偽) is produced by activated macrophages and T cells, transmembrane and secretory. TNF- 偽, an important cytokine, has a wide range of biological activities, including the regulation of hematopoiesis, immunity and inflammation. The effects of TNF- 偽 on blood vessels and coagulation and on various organs. Early studies have found that TNF- 偽 can induce necrosis or apoptosis of tumor cells, and later it was found that TNF- 偽 mediates inflammation. Major cytokines associated with cellular immune response and tumor immunity. Known diseases associated with TNF- 偽 include: AIDS-, anemia, autoimmune diseases, tumors, hemorrhagic shock, transplant rejection, tuberculosis, leukemia, diabetes, Rheumatoid arthritis and so on. TNF- 偽 is involved in the occurrence and development of many diseases, so at different levels of blocking the role of TNF- 偽, It is possible to treat TNF- 偽 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- 偽 gene sequence. The aim of this experiment was to control the secretion of TNF- 偽 by inhibiting the target gene of TNF- 偽 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.
【學(xué)位授予單位】：中國醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2005
【分類號(hào)】：R392

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