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目的:通过筛选原发性肝癌血清蛋白结合蛋白,探究肝癌的分子发病机制.方法:应用噬菌体展示技术,以肝癌患者的血清作为固相筛选分子,对T7 Select噬菌体人肝细胞cDNA文库进行5轮“吸附-洗脱-扩增”的筛选过程,经噬斑的聚合酶链式反应(PCR)扩增后,对阳性PCR产物直接进行序列测定和同源性分析.结果:噬菌体经富集后,随机挑取48个噬斑进行PCR扩增,得到5种大小不同的PCR片断,序列测定后经序列同源性分析,结果发现与肝癌患者血清蛋白结合的蛋白有以下5种:人核仁螺旋体磷酸化蛋白NOLC1,人高度迁移基核小体结合域1(HMGN1),人依赖ATP的DNA连接酶1(LIG1),人小EDRK-丰富因子2(SERF2)和A-kinase anchor protein 9 isoform.结论:用噬菌体人肝cDNA文库筛选得到了肝癌血清蛋白结合蛋白,为进一步阐明肝癌的发生及发病机制奠定了基础.
Objective: To explore the molecular pathogenesis of primary hepatocellular carcinoma by screening serum protein-binding protein of primary hepatocarcinoma.Methods: The phage display technique was used to screen the T7 Select phage human hepatocyte cDNA library with 5 cycles After the amplification by plaque polymerase chain reaction (PCR), the positive PCR products were directly sequenced and homology analysis.Results: Phage rich After collection, 48 plaques were randomly selected for PCR amplification. Five PCR fragments of different sizes were obtained. Sequence sequence analysis revealed that there were five kinds of proteins that were associated with serum protein of liver cancer patients Nucleocapsid Phosphatase NOLC1, human highly migrated nucleosome binding domain 1 (HMGN1), human DNA dependent ligase 1 (LIG1), small human EDRK-rich factor 2 (SERF2), and A-kinase anchor protein 9 isoform.Conclusion: The serum protein-binding protein of hepatocellular carcinoma was screened from the phage human liver cDNA library, which laid the foundation for further clarifying the occurrence and pathogenesis of hepatocellular carcinoma.