本文選題：染色質(zhì)隔離子　切入點：hMMP-12　出處：《新疆醫(yī)科大學》2006年碩士論文　論文類型：學位論文

【摘要】：目的：(1)構建巨噬細胞特異表達且?guī)в腥旧|(zhì)隔離子的人基質(zhì)金屬蛋白酶-12(hMMP-12)轉基因表達載體；(2)構建hMMP-12轉基因兔動物模型。方法：(1)通過RT-PCR方法擴增hMMP-12基因，產(chǎn)物亞克隆到pGEM-T載體，經(jīng)測序準確無誤后命名為pGEM-T-hMMP-12；(2)利用pJC13(backbone：pGEM4Z)，構建帶有染色質(zhì)隔離子和清道夫受體A(scavenger receptor A，SR-A)增強子、啟動子和人生長激素尾(hGH tail)的亞克隆pJC13-SR；(3)利用pGEM-T-hMMP-12和pJC13-SR重組質(zhì)粒，構建巨噬細胞特異性的轉基因表達載體pJC13-SR-hMMP-12；(4)pJC13-SR-hMMP-12表達載體線性化，顯微注射家兔受精卵制備轉基因兔，并進行PCR擴增及Southern Blot轉基因效果分析鑒定。結果：(1)通過DNA測序和限制性核酸內(nèi)切酶酶切鑒定證實，hMMP-12成功克隆到所需表達載體中；(2)轉基因后獲得仔兔36只，以PCR檢測結果計算，轉基因總效率為1.41％～3.35％，整合率為20％～50％；以Southern Blot檢測結果計算，轉基因總效率為0.47％～2.23％，整合率為13.33％～16.67％。結論：轉基因表達載體的成功構建，，為hMMP-12轉基因兔動物模型的構建和研究MMP-12在動脈粥樣硬化的形成和發(fā)展中的作用機制奠定了基礎。
[Abstract]:Objective to construct hMMP-12 transgenic rabbit model by constructing macrophage specific expression vector of human matrix metalloproteinase-12hMMP-12 with chromatin isolator. Methods the hMMP-12 gene was amplified by RT-PCR and the product was subcloned into pGEM-T vector. Using pJC13 backbone: pGEM4ZN, we constructed the enhancer with chromatin isolon and scavenger receptor Ascavenger receptor (SR-A3), the promoter and human growth hormone htail GH (htail GH) subclone pJC13-SRA3) using pGEM-T-hMMP-12 and pJC13-SR recombinant plasmids. Macrophage specific expression vector pJC13-SR-hMMP-12 was constructed to linearize the expression vector pJC13-SR-hMMP-12. The transgenic rabbits were prepared by microinjection of rabbit fertilized eggs. PCR amplification and analysis of Southern Blot transgenic effect were performed. Results: 1) DNA sequencing and restriction endonuclease digestion confirmed that hMMP-12 was successfully cloned into the required expression vector, and 36 rabbits were obtained. The results of PCR analysis were used to calculate the results of PCR detection. The total efficiency of transgenic gene was 1.41 ~ 3.35 and the integration rate was 20 / 50. Based on the results of Southern Blot test, the total efficiency of transgenic gene was 0.472.23, and the integration rate was 13.33 ~ 16.67.Conclusion: the successful construction of transgenic expression vector, It provides a basis for the construction of hMMP-12 transgenic rabbit model and the study of the role of MMP-12 in the formation and development of atherosclerosis.
【學位授予單位】：新疆醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2006
【分類號】：R346

【參考文獻】

相關期刊論文 前1條

1 郭愛桃,韋立新,石懷銀,李向紅,游聯(lián)璧;基質(zhì)金屬蛋白酶1與冠狀動脈粥樣硬化斑塊破裂的關系[J];中華病理學雜志;2000年04期

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