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目的运用生物信息学方法预测人Sox10基因启动子区甲基化情况并进行实验验证。方法从GenBank获取人Sox10基因序列,利用Methyprimer 11.0软件、Li lab、CpG Island Searcher预测其启动子区CpG岛。进一步运用甲基化特异性PCR(methylation specific PCR,MSP)和重亚硫酸盐测序(bisulfite sequencing PCR,BSP)方法检测人Sox10基因启动子区CpG岛在多形性胶质母细胞瘤和瘤周组织中的甲基化情况。结果经过生物信息学分析发现,人Sox10基因启动子区存在2个CpG岛,分别为165(1 241～1 405)bp和105(1 478～1 582)bp。针对这两个CpG岛设计引物行MSP和BSP实验,研究发现,在人多形性胶质母细胞瘤和瘤周组织中Sox10基因启动子区CpG岛均出现部分甲基化,其中在多形性胶质母细胞瘤中的甲基化率为90.5%,在瘤周组织中的甲基化率为28.5%。结论本研究证实了人Sox10基因启动子区CpG岛在多形性胶质母细胞瘤中出现高甲基化,提示其表达下调可能受甲基化调节。
Objective To predict the methylation of human Sox10 promoter region by bioinformatics and to verify the results. Methods The human Sox10 gene sequence was obtained from GenBank. The CpG islands of CpG islands were predicted by Methyprimer 11.0 software, Li lab and CpG Island Searcher. Furthermore, methylation-specific PCR (MSP) and bisulfite sequencing PCR (BSP) were used to detect the expression of CpG island in human glioma cell line Glioblastoma and tumor Methylation in the tissue. Results After bioinformatics analysis, there were two CpG islands in the human Sox10 gene promoter, which were 165 (1 241 ~ 1 405) bp and 105 (1 478 ~ 1 582) bp, respectively. For these two CpG island design primer MSP and BSP experiments found that in human glioblastoma multiforme and tumor tissue in the promoter region CpG island Sox10 partial methylation, which polymorphism The rate of methylation in glioblastoma was 90.5% and the methylation rate in tumor tissues was 28.5%. Conclusion Our study confirmed that CpG island of human Sox10 gene promoter is hypermethylated in glioblastoma multiforme, suggesting that its downregulation might be regulated by methylation.