关键词: 肿瘤;mRNA差异显示法;银染;基因克隆 中图号:R735.7 文献标识码:A

    摘 要:目的 建立银染mRNA差异显示方法，筛选并克隆肝癌相关基因. 方法 以建系的人肝癌细胞HepG2和正常的肝细胞L02的总RNA为模板，用5'-dT11 G锚定引物和8条随机引物(AP1～AP8)组合进行RT-PCR扩增，PCR产物经60g·L-1 尿素变性聚丙烯酰胺凝胶电泳分离后，凝胶银盐染色显示差异的DNA片段. 结果 建立了快速敏感的银染mRNA差异显示方法，分离并克隆了一些肝癌相关基因. 结论 建立的银染mRNA差异显示法简单、有效，在差异表达基因的筛选中有较高的实用价值.

    Silver┐staining mRNA differential display method and cloning of tumor related genes in HepG2cell line

    WANG Xin，HUANG Yu-Xin，WEN Qin-Sheng，WANG Qing-Li

    Department of Gastroenterology，Tangdu Hospital，Fourth Military Medical University，Xi'an710038，China

    Keywords:tumor;mRNA differential display;silver stain-ing;cloning of gene

    Abstract:AIM To develop mRNA differential display poly-merize chain reaction(DD-PCR)method with silver staining and to clone the liver cancer related genes in HepG2cell line.METHODS Total RNA was extracted from the HepG2cells and L02cells.Their first strains of cDNA were produced by reverse transcription(RT).The cDNA were amplified by PCR using an anchored primer5'dT11G combined with eight arbitrary primers respectively.The products of PCR were an-alyzed on a denaturing60g·L-1 polyacrylamide gel.The differential DNA bands on gel were seen by silver-staining. ......