摘要
目的 建立银染 m RNA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 Hep G2和正常的肝细胞 L0 2的总 RNA为模板 ,用 5 - d T1 1 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 6 0 g· 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m RNA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 m RNA差异显示法简单、有效 。
AIM To develop mRNA differential display polymerize chain reaction (DD PCR) method with silver staining and to clone the liver cancer related genes in HepG2 cell line. METHODS Total RNA was extracted from the HepG2 cells and L02 cells. Their first strains of cDNA were produced by reverse transcription (RT). The cDNA were amplified by PCR using an anchored primer 5 dT11G combined with eight arbitrary primers respectively. The products of PCR were analyzed on a denaturing 60 g·L -1 polyacrylamide gel. The differential DNA bands on gel were seen by silver staining.RESULTS Several liver cancer related genes were isolated and identified by the sensitive silver staining mRNA differential display developed. CONCLUSION The established mRNA differential display method with silver staining is a simple, efficient means which has higher value for screening and cloning differentially expressed genes.
出处
《第四军医大学学报》
北大核心
2001年第9期843-845,共3页
Journal of the Fourth Military Medical University
关键词
肝癌
MRNA差异显示法
基因克隆
银染
tumor
mRNA differential display
silver staining
cloning of gene
