本文關(guān)鍵詞：STAT6基因RNAi表達(dá)載體的構(gòu)建及其沉默效應(yīng)觀察　出處：《重慶醫(yī)科大學(xué)》2010年碩士論文　論文類(lèi)型：學(xué)位論文

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【摘要】： 哮喘是以氣道高反應(yīng)性和慢性氣道炎癥為特征的變態(tài)反應(yīng)疾病。Th1/Th2平衡失調(diào),即Th1功能下降,Th2功能亢進(jìn)是目前比較公認(rèn)的哮喘免疫學(xué)發(fā)病機(jī)制。由于特異性調(diào)控Th0細(xì)胞向Th2分化,STAT6基因成為治療哮喘的靶基因。目前尚未報(bào)道STAT6基因的高效特異性的SiRNA序列,因此本研究選擇STAT6基因作為RNA干擾(RNA interference,RNAi)的靶點(diǎn),構(gòu)建STAT6干擾質(zhì)粒,為哮喘的治療研究提供新思路。 目的:構(gòu)建針對(duì)STAT6基因的干擾質(zhì)粒,并觀察其對(duì)STAT6蛋白表達(dá)的抑制效果。 方法:通過(guò)體外轉(zhuǎn)錄合成shRNA,與pIRES2  EGFP  STAT6基因表達(dá)質(zhì)粒共同轉(zhuǎn)染COS7細(xì)胞,在熒光顯微鏡下觀察干擾效果,同時(shí)從蛋白水平對(duì)干擾質(zhì)粒的抑制效果進(jìn)行驗(yàn)證。 結(jié)果:將pIRES2  EGFP  STAT6重組載體轉(zhuǎn)染COS7細(xì)胞后,結(jié)果顯示STAT6蛋白可以在細(xì)胞中高水平表達(dá)。與正常對(duì)照組相比,pGenesil3-STAT61和pGenesil3-STAT62對(duì)STAT6蛋白的抑制率分別是74.3%和85.8%,其中pGenesil3-STAT62的抑制作用更為顯著。 結(jié)論:成功構(gòu)建并篩選到具有顯著干擾效率的STAT6干擾質(zhì)粒,為進(jìn)一步研究STAT6在哮喘發(fā)病機(jī)制中的作用,同時(shí)為哮喘治療提供新的思路。
[Abstract]:Asthma is an allergic disease characterized by airway hyperresponsiveness and chronic airway inflammation. Th1 / Th2 imbalance is called the decline of Th1 function. Hyperfunction of Th2 is recognized as the immunological mechanism of asthma, and the differentiation of Th0 cells into Th2 is regulated by specific regulation. STAT6 gene has become the target gene for the treatment of asthma. At present, the highly efficient and specific SiRNA sequence of STAT6 gene has not been reported. In this study, STAT6 gene was selected as the target of RNA interference RNA interference to construct STAT6 interference plasmid. To provide a new idea for the treatment of asthma. Aim: to construct the interference plasmid for STAT6 gene and observe its inhibitory effect on the expression of STAT6 protein. Methods: shRNAs were synthesized by transcription in vitro and transfected into COS7 cells with pIRES2 EGFP STAT6 gene expression plasmid. The interference effect was observed under fluorescence microscope and the inhibition effect of interference plasmid was verified from protein level. Results: the recombinant vector of pIRES2 EGFP STAT6 was transfected into COS7 cells. The results showed that STAT6 protein could be expressed at a high level in the cells compared with the normal control group. The inhibitory rates of pGenesil3-STAT61 and pGenesil3-STAT62 on STAT6 protein were 74.3% and 85.8%, respectively. The inhibitory effect of pGenesil3-STAT62 was more significant. Conclusion: STAT6 interference plasmids with significant interference efficiency were successfully constructed and screened to further study the role of STAT6 in the pathogenesis of asthma and to provide new ideas for the treatment of asthma.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R346

【引證文獻(xiàn)】

相關(guān)博士學(xué)位論文 前1條

1 應(yīng)林燕;CTLA4Ig修飾的樹(shù)突狀細(xì)胞對(duì)哮喘小鼠氣道炎癥及Th失衡的干預(yù)作用的實(shí)驗(yàn)研究[D];重慶醫(yī)科大學(xué);2011年

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本文編號(hào)：1377172