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目的 克隆新的肝癌相关基因 ,探索肝癌发生的分子机制。方法 采用DDPCR技术 ,分析原发性肝癌和癌旁肝组织间基因表达差异情况 ,获得差异表达基因片段 ,Northern杂交证实该基因片段能与原标本印迹重现 ;用此基因片段作为探针 ,通过筛选胎盘cDNA文库而获得新基因全长 ;利用GST融合蛋白制备新基因C端多克隆抗体 ,Western杂交验证抗体的特异性和目的基因的表达。结果 成功克隆了一个富含脯氨酸的肝癌新基因cDNA全长 ,并制备出新基因多克隆抗体。结论 我们克隆的这一新的肝癌差异表达基因和制备的该基因C端多克隆抗体为肝癌的基因治疗打下了基础。
Objective To clone new liver cancer related genes and explore the molecular mechanism of liver cancer. Methods DDPCR technique was used to analyze the gene expression differences between primary liver cancer and adjacent liver tissue. Differentially expressed gene fragments were obtained. Northern blotting confirmed that the gene fragment could be reproduced with the original sample. This gene fragment was used as a probe and passed. The placenta cDNA library was screened to obtain the full length of the new gene; a polyclonal antibody at the C-terminus of the new gene was prepared using the GST fusion protein, and Western blotting verified the specificity of the antibody and the expression of the target gene. Results The full-length cDNA of a novel hepatocarcinoma gene rich in proline was successfully cloned and a new polyclonal antibody was prepared. Conclusion The new differentially expressed hepatocellular carcinoma gene and the polyclonal antibody at the C-terminus of this gene have laid a foundation for the gene therapy of liver cancer.