染色體12q14.2區(qū)域重測序篩選卵巢癌易感性SNP位點及其分子機制研究
發(fā)布時間：2018-07-20 21:00
【摘要】：目的卵巢癌是一種常見婦科腫瘤,由于缺乏早期相關(guān)明顯癥狀而常被喻為“沉默殺手”。其是一種復(fù)雜的多基因疾病,遺傳因素發(fā)揮著重要作用。為了發(fā)現(xiàn)中國人特有的遺傳易感信息,課題組前期開展的中國漢族女性上皮源性卵巢癌GWAS研究發(fā)現(xiàn)了12q14.2區(qū)的rs11175194位點與國人卵巢癌發(fā)病風(fēng)險相關(guān),其不同基因型與基因SRGAP1表達相關(guān),基因SRGAP1高表達組卵巢癌患者預(yù)后差,提示此高連鎖不平衡易感區(qū)域重要,可能成為卵巢癌高危篩查、診斷和治療靶點。本研究擬對該區(qū)域進行重測序,對該區(qū)域進行精細作圖,篩選病例對照有差異的SNP位點,分析SNP基因型與環(huán)境因素的交互作用,預(yù)測該區(qū)域差異SNP卵巢癌易感性位點的功能,進行功能探索,同時結(jié)合美國TCGA數(shù)據(jù)進行驗證,為闡明卵巢癌的病因和發(fā)生發(fā)展機制,篩選高危易感人群,開展早期診斷及預(yù)后預(yù)測提供新依據(jù),為提高卵巢癌早診率降低死亡率提供參考。方法基于Hapmap里中國及日本人群的數(shù)據(jù),利用Haploiew軟件,對中國漢族人群卵巢癌GWAS研究發(fā)現(xiàn)的易感性位點rs11175194進行連鎖不平衡分析,得到該位點所在的連鎖不平衡區(qū)域。采用高通量二代測序方法,對192例正常對照人群中該區(qū)域進行重測序,分析中國漢族女性該區(qū)域中已知和未知變異位點,剔除MAF0.05的位點,篩選與rs11175194高度連鎖的位點進行病例對照研究,找出病例對照中SNP頻率有差異的位點,利用生物信息學(xué)方法進行功能預(yù)測。分析有功能SNP位點不同基因型與其所在基因表達量之間的關(guān)系,同時利用TCGA公共數(shù)據(jù)庫中卵巢癌患者基因表達數(shù)據(jù)信息以及相應(yīng)體外細胞實驗進行驗證。結(jié)果分析得到中國漢人卵巢癌GWAS研究發(fā)現(xiàn)的易感性SNP位點rs11175194所在的高度連鎖不平衡區(qū)域(linkage disequilibrium block,LD block)64215629-64268005之間全長52kb。經(jīng)過對正常對照人群該區(qū)域重測序,篩選與rs11175194高度連鎖并且病例對照頻率差異有意義的SNP位點rs11175195和rs67799338。這兩個位點均位于SRGAP1基因的第一號內(nèi)含子區(qū)域,生物信息學(xué)方法預(yù)測結(jié)果顯示,SNP位點rs67799338所在序列存在人源性轉(zhuǎn)錄因子Ref1、AP1、SP1結(jié)合位點。SNP位點rs67799338不同基因型與其所在基因SRGAP1的mRNA表達相關(guān),C基因型的表達量高于T基因型。TCGA公共數(shù)據(jù)庫卵巢癌患者基因表達數(shù)據(jù)分析結(jié)果顯示,轉(zhuǎn)錄因子Ref1、AP1、SP1的表達量與基因SRGAP1的表達量呈正相關(guān),其中轉(zhuǎn)錄因子SP1的表達量與基因SRGAP1表達量相關(guān)性最強,r=0.607,p0.001。CHIP實驗結(jié)果顯示,轉(zhuǎn)錄因子SP1可以與rs67799338所在區(qū)域結(jié)合。過表達SRGAP1可以促進SKOV3細胞進入S期。過表達SRGAP1可以增加總Rho含量,SRGAP1高表達的卵巢癌患者預(yù)后差。結(jié)論位于SRGAP1基因第一號內(nèi)含子上卵巢癌易感性SNP位點rs11175195、rs67799338,其不同基因型均影響所在基因SRGAP1的表達量。以上兩個SNP位點為C基因型時其所在基因SRGAP1的mRNA表達量高于T基因型。SNP位點rs67799338所在區(qū)域具有增強子作用,并與轉(zhuǎn)錄因子SP1結(jié)合。SRGAP1高表達的卵巢癌患者生存時間短,預(yù)后差。SRGAP1有可能成為卵巢癌預(yù)后預(yù)測的新靶標。
[Abstract]:Objective ovarian cancer is a common gynecologic tumor, which is often referred to as "silent killer" due to the lack of early related obvious symptoms. It is a complex polygenic disease, and the genetic factor plays an important role. In order to find the unique genetic susceptibility information of Chinese people, the Chinese Han women's epithelial ovarian cancer GWA has been developed in the earlier period. The S study found that the rs11175194 loci in the 12q14.2 region are associated with the risk of ovarian cancer in the country. The different genotypes are associated with the gene expression of SRGAP1, and the prognosis of the ovarian cancer patients with the high expression of SRGAP1 is poor, suggesting that the high linkage unbalance susceptible region is important and may be a high risk screening, diagnosis and therapeutic target for ovarian cancer. This study is to be used in this study. The region was re sequenced, the region was carefully plotted, the SNP loci with different case control were screened, the interaction between the SNP genotype and the environmental factors was analyzed, the function of the SNP ovarian cancer susceptibility loci was predicted, and the functional exploration was carried out. At the same time, the TCGA data of the United States were used to verify the etiology and occurrence of ovarian cancer. It provides a new basis for early diagnosis and prognosis prediction of high-risk and susceptible population, and provides a reference for improving the early diagnosis rate of ovarian cancer. Methods based on the data of Chinese and Japanese people in Hapmap, the Haploiew software is used to chain the susceptible locus rs11175194 found in the GWAS study of Chinese Han population. The linkage disequilibrium region of the loci was obtained by unbalanced analysis. A high throughput two generation sequencing method was used to re sequence the region in 192 normal controls. The known and unknown mutation sites in the Chinese Han women were analyzed, the MAF0.05 loci were eliminated and the highly linked rs11175194 loci were screened for case control research. To find out the loci with different frequencies of SNP in the case control and use the Bioinformatics Method to predict the function, the relationship between the different genotypes of the functional SNP loci and the expression of their genes is analyzed, and the data of the gene expression data of the ovarian cancer patients in the TCGA public database and the corresponding in vitro cell experiment are verified. The results were obtained from the high linkage disequilibrium region (linkage disequilibrium block, LD block) 64215629-64268005 of the susceptibility SNP locus (linkage disequilibrium block, LD block) 64215629-64268005, which was found in the Chinese Han Chinese ovarian cancer study (linkage disequilibrium block, LD block). The total length of 52kb. was re sequenced in the normal control population, screening with rs11175194 highly linked and case control frequency difference. The two loci of SNP loci rs11175195 and rs67799338. are located in the first intron of the SRGAP1 gene. The results of bioinformatics methods show that the sequence of SNP loci rs67799338 exists the human source transcription factor Ref1, AP1, SP1 binding site.SNP locus rs67799338 different genotypes and their genes SRGAP1. The expression of A was related, the expression of C genotype was higher than that of the T genotype.TCGA public database of ovarian cancer patients. The expression of Ref1, AP1, SP1 was positively correlated with the expression of gene SRGAP1, and the expression of the transcription factor SP1 was strongest with the expression of gene SRGAP1, r=0.607, p0.001.CHIP experiment The results showed that the transcription factor SP1 could be combined with the region of rs67799338. Overexpression of SRGAP1 could promote the entry of SKOV3 cells into the S phase. The overexpression of SRGAP1 could increase the total Rho content, and the prognosis of ovarian cancer patients with high expression of SRGAP1 was poor. Conclusion the SNP locus of the susceptibility to ovarian cancer in the first intron No. 1 of the SRGAP1 gene was rs11175195, rs67799338, it was not. The same genotype affects the expression of the gene SRGAP1. When the two SNP loci are C genotype, the mRNA expression of the gene SRGAP1 is higher than that of the T genotype.SNP locus rs67799338, and the survival time of the ovarian cancer patients with the high expression of the transcription factor SP1 binding.SRGAP1 is short, and the poor prognosis is possible. It is a new target for prognosis prediction of ovarian cancer.
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R737.31
本文編號：2134761
[Abstract]:Objective ovarian cancer is a common gynecologic tumor, which is often referred to as "silent killer" due to the lack of early related obvious symptoms. It is a complex polygenic disease, and the genetic factor plays an important role. In order to find the unique genetic susceptibility information of Chinese people, the Chinese Han women's epithelial ovarian cancer GWA has been developed in the earlier period. The S study found that the rs11175194 loci in the 12q14.2 region are associated with the risk of ovarian cancer in the country. The different genotypes are associated with the gene expression of SRGAP1, and the prognosis of the ovarian cancer patients with the high expression of SRGAP1 is poor, suggesting that the high linkage unbalance susceptible region is important and may be a high risk screening, diagnosis and therapeutic target for ovarian cancer. This study is to be used in this study. The region was re sequenced, the region was carefully plotted, the SNP loci with different case control were screened, the interaction between the SNP genotype and the environmental factors was analyzed, the function of the SNP ovarian cancer susceptibility loci was predicted, and the functional exploration was carried out. At the same time, the TCGA data of the United States were used to verify the etiology and occurrence of ovarian cancer. It provides a new basis for early diagnosis and prognosis prediction of high-risk and susceptible population, and provides a reference for improving the early diagnosis rate of ovarian cancer. Methods based on the data of Chinese and Japanese people in Hapmap, the Haploiew software is used to chain the susceptible locus rs11175194 found in the GWAS study of Chinese Han population. The linkage disequilibrium region of the loci was obtained by unbalanced analysis. A high throughput two generation sequencing method was used to re sequence the region in 192 normal controls. The known and unknown mutation sites in the Chinese Han women were analyzed, the MAF0.05 loci were eliminated and the highly linked rs11175194 loci were screened for case control research. To find out the loci with different frequencies of SNP in the case control and use the Bioinformatics Method to predict the function, the relationship between the different genotypes of the functional SNP loci and the expression of their genes is analyzed, and the data of the gene expression data of the ovarian cancer patients in the TCGA public database and the corresponding in vitro cell experiment are verified. The results were obtained from the high linkage disequilibrium region (linkage disequilibrium block, LD block) 64215629-64268005 of the susceptibility SNP locus (linkage disequilibrium block, LD block) 64215629-64268005, which was found in the Chinese Han Chinese ovarian cancer study (linkage disequilibrium block, LD block). The total length of 52kb. was re sequenced in the normal control population, screening with rs11175194 highly linked and case control frequency difference. The two loci of SNP loci rs11175195 and rs67799338. are located in the first intron of the SRGAP1 gene. The results of bioinformatics methods show that the sequence of SNP loci rs67799338 exists the human source transcription factor Ref1, AP1, SP1 binding site.SNP locus rs67799338 different genotypes and their genes SRGAP1. The expression of A was related, the expression of C genotype was higher than that of the T genotype.TCGA public database of ovarian cancer patients. The expression of Ref1, AP1, SP1 was positively correlated with the expression of gene SRGAP1, and the expression of the transcription factor SP1 was strongest with the expression of gene SRGAP1, r=0.607, p0.001.CHIP experiment The results showed that the transcription factor SP1 could be combined with the region of rs67799338. Overexpression of SRGAP1 could promote the entry of SKOV3 cells into the S phase. The overexpression of SRGAP1 could increase the total Rho content, and the prognosis of ovarian cancer patients with high expression of SRGAP1 was poor. Conclusion the SNP locus of the susceptibility to ovarian cancer in the first intron No. 1 of the SRGAP1 gene was rs11175195, rs67799338, it was not. The same genotype affects the expression of the gene SRGAP1. When the two SNP loci are C genotype, the mRNA expression of the gene SRGAP1 is higher than that of the T genotype.SNP locus rs67799338, and the survival time of the ovarian cancer patients with the high expression of the transcription factor SP1 binding.SRGAP1 is short, and the poor prognosis is possible. It is a new target for prognosis prediction of ovarian cancer.
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R737.31
【參考文獻】
相關(guān)期刊論文 前1條
1 郭俊;李小燕;蔡倫;王綠婭;杜杰;;醫(yī)學(xué)研究生"高通量測序技術(shù)"應(yīng)用能力的培養(yǎng)[J];現(xiàn)代生物醫(yī)學(xué)進展;2016年31期
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