dna脱甲基化对整合至rdna区源基因hfⅷ表达量的影响-遗传学专业毕业论文.docxVIP

硕士学位论文 中文摘要 2’一deoxycytidine处理72小时和120小时的细胞，hFⅧ的表达量分 别是未加脱甲基化药物处理对照组的5倍和10倍。重亚硫酸盐测序 PCR的结果显示目的基因CMV启动子区未发现有甲基化位点。 结论：DNA甲基化修饰对定点整合目的基因的表达有较大影响， 但是这种影响可能并非由目的基因启动子区异常甲基化引起的。我们 推测rDNA区脱甲基化引起染色质结构变化可能是造成这一结果的原 因。 关键词DNA甲基化，人凝血因子Ⅷ，ELISA，pHrneo Ⅱ  硕士学位论文 英文摘要 ABSTRACT As a kind of gene expression regulation mechanism,DNA methylation play all important role in many biology processes，including embryonic development，genetic imprinting，cell differentiation， tumorigenesis and SO on．pHrneo，a human ribosomal DNA—targeting vector developed by OI／17 group，Can target exogenous gene to human ribosomal DNA locus．The exogenous genes carded by pHrneo could stably express in some cell lines，such as HL7702(human hepatocyte) and HTl080(human fibrosarcoma)．But the advanced efficiency of expression is still OUr aim．The factors affecting interested site-directed integration gene expression may involve copy numbers of interested gene， DNA methylation modification，histone acetylation modification and SO on．In this study,our focus is on the relation between interested site-directed integration gene expression and DNA methlation Objects：In this research，we investigated the relation between interested site—directed integration gene expression and DNA methlation modification by comparing human coagulation factor FⅧexpression level of site—directed integration cell between demethylation agent 5一aza一 2-deoxycytidine treatment group and control group，and the methylation pattem of interested gene promoter． Methods：Three concentrations of 5一aza一2-deoxycitidine(5“M， m  硕士学位论文 英文摘要 l O-tM，20txM)were respectively applied to treat site—directed integration cell pHrneoSK39-25 for 72h and 1 20h．Wild site—directed integration cell pHrneoSK39—25，HTl080 cell treated by 5-aza-2-deox