本文選題：單核苷酸多態(tài)性　切入點：冠心病　出處：《延邊大學(xué)》2015年碩士論文

【摘要】：目的：在中國延邊地區(qū)人群中尋找一個冠心病(CAD)易感性標(biāo)記物。方法：本研究應(yīng)用計算機生物信息學(xué)預(yù)測軟件和NCBI-SNP數(shù)據(jù)庫,從脂聯(lián)素及其受體基因中篩選出和miRNA結(jié)合能差異(|△△G|)大于5.27 kcal/mol的6個野生型和變異型等位基因。采用聚合酶鏈反應(yīng)-限制性片段長度多態(tài)法(PCR-RFLP)方法對6個野生型和變異型等位基因進行基因分型,比較基因型的分布頻率,觀察與冠心病易感性的相關(guān)性。應(yīng)用PCR擴增3UTR基因組DNA,利用限制性內(nèi)切酶切斷PCR產(chǎn)物及pGL3系列的質(zhì)粒,利用T4DNA連接酶連接兩種DNA,制備pGL3-3'UTR系列的質(zhì)粒,在E.coli中大量培養(yǎng)后與miRNA表達質(zhì)粒一起轉(zhuǎn)染給人平滑肌細胞株培養(yǎng),進行熒光素酶檢測,用含海腎螢光素酶(Renilla luciferase)的pRL-SV40質(zhì)粒作為內(nèi)部對照物,利用雙熒光素酶基因檢測系統(tǒng)測定啟動子活性.,利用Trizol法提取不同基因型RNA后反轉(zhuǎn)錄為cDNA,利用qRT-PCR方法比較不同基因型的mRNA表達量。采用Western-blot法檢測結(jié)果顯示不同基因型的ADIPOR1蛋白表達量。結(jié)果：1.ADIPORl-rs7539542C等位基因冠心病發(fā)病率顯著高于其他等位基因。2.通過用熒光素酶檢測測定啟動子活性,結(jié)果顯示只有ADIPORl-rs7539542C有顯著性統(tǒng)計學(xué)意義,p為0.03。3.qRT-PCR方法顯示,rs7539542G時mRNA表達量顯著低于rs7539542C,具有統(tǒng)計學(xué)意義,p為0.02。4. Western-blot法檢測結(jié)果顯示,rs7539542G時mRNA表達量顯著低于rs7539542C,具有統(tǒng)計學(xué)意義,p為0.01。結(jié)論：本研究結(jié)果顯示延邊地區(qū)人群冠心病易感基因,ADIPORl-rs7539542C可作為延邊地區(qū)預(yù)測冠心病易感性標(biāo)志物。
[Abstract]:Objective: to search for a susceptible marker of coronary artery disease (CAD) in the population of Yanbian area, China. Methods: the computer bioinformatics prediction software and NCBI-SNP database were used in this study. Six wild and variant alleles with miRNA binding energy (G) greater than 5.27 kcal/mol were screened from adiponectin and its receptor genes. Genetic and variant alleles were genotyped. To compare the frequency of genotype distribution and to observe the correlation between genotype distribution and susceptibility to coronary heart disease (CHD), PCR was used to amplify 3UTR genomic DNA, restriction endonuclease was used to cut off PCR products and plasmids of pGL3 series, T4DNA ligase was used to connect the two kinds of plasmids, and plasmids of pGL3-3'UTR series were prepared. Human smooth muscle cell lines were transfected with miRNA expression plasmid in large quantities in E.coli. Luciferase was detected by luciferase assay. The pRL-SV40 plasmid containing luciferase was used as internal control. The promoter activity was determined by double luciferase gene detection system, the RNA of different genotypes was extracted by Trizol method, and the mRNA expression of different genotypes was compared by qRT-PCR method. The results of Western-blot detection showed different bases. Results: 1. The incidence of coronary heart disease of ADIPORl-rs7539542C allele was significantly higher than that of other alleles .2.The promoter activity was determined by luciferase assay. The results showed that the expression of mRNA was significantly lower than that of rs7539542G by 0.03.3.qRT-PCR method, and was significantly lower than that of rs7539542G by 0.03.3.qRT-PCR method. The results of Western-blot assay showed that the expression of mRNA was significantly lower than that of rs7539542G, and the results of Western-blot showed that the expression of mRNA was significantly lower than that of rs7539542G. Conclusion: the results showed that ADIPORl-rs7539542C could be used as a marker to predict the susceptibility of coronary heart disease in Yanbian area.
【學(xué)位授予單位】：延邊大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R541.4

【參考文獻】

相關(guān)期刊論文 前3條

1 陳芳;蘇穎;李曉霞;武海亮;王潔;楊銳英;;脂聯(lián)素水平及基因多態(tài)性與代謝綜合征的關(guān)系[J];臨床心血管病雜志;2011年11期

2 茹松偉;申衛(wèi)紅;楊鵬程;趙屹;邵啟祥;;microRNA靶基因預(yù)測算法研究概況及發(fā)展趨勢[J];生命科學(xué);2007年05期

3 李曉燕;郭文靜;張紅明;劉科衛(wèi);韓淑芳;;急性冠狀動脈綜合征患者血漿脂聯(lián)素的變化[J];中華臨床醫(yī)師雜志(電子版);2013年08期

，

本文編號：1660195