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目的:筛选和克隆肝癌和正常肝组织差异表达的基因片段,以探讨肝癌的发病机制.方法:应用mRNA差异显示技术(mRNA-DD),比较肝癌及正常肝组织基因表达的差异,对获得的差异基因片段进行克隆、测序,测序结果提交GenBank数据库中进行同源性分析,并对其中一条有明显差异的基因用半定量RT-PCR分析该基因在正常肝组织及肝癌组织中的表达情况.结果:凝胶扫描显示在500-600 bp处有一条差异片段,差异条带经酶切图谱分析证实重组质粒是目的克隆,将片段测序结果与GenBank数据库进行同源性比较发现该片段与已知基因EEG1高度同源(99%).RT-PCR结果显示在肝癌组织中EEG1 mRNA的表达水平明显低于正常肝癌组织(P=0.001).结论:EEG1基因在肝癌组织中的表达明显低于正常肝组织,提示EEG1基因的表达下调可能与肝癌的发病有关.
OBJECTIVE: To screen and clone differentially expressed genes in hepatocellular carcinoma and normal liver tissues to explore the pathogenesis of hepatocellular carcinoma.Methods: mRNA expression differences (mRNA-DD) were compared between hepatocellular carcinoma and normal liver tissue, Gene fragments were cloned, sequenced and sequenced. The results were submitted to GenBank for homology analysis, and one of the genes with significant difference was analyzed by semi-quantitative RT-PCR in normal liver tissue and hepatocellular carcinoma. Results: The gel scan showed a different fragment at 500-600 bp. The different bands were confirmed by restriction enzyme digestion analysis. The comparison of the sequence of the fragment with the GenBank database showed that the fragment was homologous with the known The gene EEG1 was highly homologous (99%). The results of RT-PCR showed that the expression level of EEG1 mRNA in HCC tissues was significantly lower than that in normal liver cancer tissues (P = 0.001) .Conclusion: The expression of EEG1 gene in HCC tissues was significantly lower than that in normal liver tissues Liver tissue, suggesting that the expression of EEG1 gene may be related to the incidence of liver cancer.