本文選題：左向右分流型先天性心臟病　切入點(diǎn)：心肌纖維化　出處：《中南大學(xué)》2012年碩士論文

【摘要】：目的通過(guò)檢測(cè)左向右分流型先天性心臟病患兒血細(xì)胞中TGF-β1基因多態(tài)性位點(diǎn)+869T/C表達(dá)水平,探討TGF-β1與左向右分流型先天性心臟病心肌纖維化的相關(guān)性,以及基因位點(diǎn)+869T/C基因多態(tài)性在心肌纖維化過(guò)程中的意義。 方法隨機(jī)選擇同期行手術(shù)治療或心臟彩超證實(shí)有左向右分流型先天性心臟病(室間隔缺損、房間隔缺損、動(dòng)脈導(dǎo)管未閉)患兒43例,其中男26例、女17例,年齡0.28～8歲,平均1.43±2.65歲。匹配正常健康兒童35例,其中男23例、女12例,年齡0.39～7歲,平均2.51±2.24歲。獲得其父母知情同意后晨起空腹采靜脈血,提取血細(xì)胞中的DNA,通過(guò)建立TGF-β1/β-actin實(shí)時(shí)熒光定量PCR檢測(cè)方法對(duì)臨床樣本進(jìn)行檢測(cè),比較兩組間兩個(gè)不同等位基因的基因拷貝數(shù)分布頻率,分析+869T/C基因拷貝數(shù)變異與心肌纖維化的關(guān)系。 結(jié)果(1)利用紫外分光光度儀對(duì)提取的總DNA進(jìn)行光度吸收測(cè)量,OD26o/OD280在1.7-1.9之間,表明DNA的純度及濃度在理想范圍內(nèi)；總DNA電泳結(jié)果顯示：240bp和137bp條帶清晰,無(wú)拖尾；灰度比大于1,表明DNA完整性好。(2) TGF-β1+869T/C和β-actin的標(biāo)準(zhǔn)曲線線性關(guān)系良好,相關(guān)系數(shù)(R2)均大于0.99。TGF-β1+869T/C標(biāo)準(zhǔn)曲線方程為:Y=-0.2964X+7.8603,R2=0.999; β-actin的標(biāo)準(zhǔn)曲線方程為：Y=-0.2888X+0.9989,R2=0.9989。(3)實(shí)驗(yàn)組和對(duì)照組+869/β-actin比值分別為0.0881±0.0397(n=43)和0.0436士0.0061(n=35),t=2.632(n=78,p0.05),差異有統(tǒng)計(jì)學(xué)意義。(4)實(shí)驗(yàn)組和對(duì)照組三種基因型(TC, TT, CC)實(shí)際值和預(yù)測(cè)值經(jīng)卡方檢驗(yàn)測(cè)得p0.05,差異無(wú)統(tǒng)計(jì)學(xué)意義。 結(jié)論(1)利用TaqMan探針實(shí)時(shí)熒光定量PCR技術(shù)證實(shí)人類基因組中TGF-β1基因在左向右分流型先天性心臟病患兒中高表達(dá)。(2)左向右分流型先心病組與正常對(duì)照組比較不同基因型TGF-β1+869基因位點(diǎn)存在T/C突變。(3)本實(shí)驗(yàn)運(yùn)用的TaqMan探針實(shí)時(shí)熒光定量PCR法具有高度的特異性和可靠性,是理想的檢測(cè)基因表達(dá)水平的方法。
[Abstract]:Objective to investigate the relationship between TGF- 尾 1 gene polymorphism and myocardial fibrosis in children with left to right shunt congenital heart disease by detecting the expression of TGF- 尾 1 gene polymorphism in blood cells of children with left to right shunt congenital heart disease.And the significance of 869T/C gene polymorphism in myocardial fibrosis.Methods Forty-three children with left to right shunt congenital heart disease (ventricular septal defect, atrial septal defect, patent ductus arteriosus) were randomly selected for surgical treatment or echocardiography, including 26 males and 17 females aged 0.288 years.The average age was 1.43 鹵2.65 years old.35 healthy children were matched, including 23 males and 12 females, aged 0.397 years, with an average age of 2.51 鹵2.24 years.After obtaining the informed consent of his parents, the venous blood was collected on an empty stomach in the morning to extract the DNA from the blood cells. The clinical samples were detected by real-time fluorescence quantitative PCR assay of TGF- 尾 1 / 尾 -actin, and the frequency of gene copy number distribution of two different alleles between the two groups was compared.To analyze the relationship between 869T/C gene copy number variation and myocardial fibrosis.The results showed that the purity and concentration of DNA were within the ideal range, and the results of total DNA electrophoresis showed that the bands of the total DNA extracted by UV spectrophotometer were clear, and the bands of 137bp and the bands of 137bp were clear and untrailing, and the absorbance of OD26o- / OD280 was between 1.7-1.9, and the results showed that the purity and concentration of DNA were in the ideal range.The grayscale ratio of TGF- 尾 1 and 尾 -actin is higher than 1, which indicates that the standard curve of DNA has good linear relationship with that of 尾 -actin and TGF- 尾 1 869T/C.The actual values and predicted values of TC, TTand CCS were measured by chi-square test (p 0.05), and the difference was not statistically significant.Conclusion: TGF- 尾 1 gene expression in human genome was confirmed by real-time fluorescence quantitative PCR with TaqMan probe in children with left to right shunt congenital heart disease.TGF- 尾 1 869 gene locus has a T / C mutation. (3) the TaqMan probe real-time quantitative PCR method used in this study is highly specific and reliable.It is an ideal method to detect gene expression level.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R725.4

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本文編號(hào)：1700865