应用代表性差异分析法筛选人癌候选抑癌基因

运用cDNA代表性差异分析法(cDNA representational difference analysis，cDNA RDA)，以正常人鼻咽上皮细胞及鼻咽癌HNE1细胞作为比较的样品来源，分离了四个在鼻咽癌中缺失的cDNA片段.以此四个片段作探针，分别进行DNA杂交、RNA杂交，结果显示,这些差异性的cDNA序列确实来自正常人鼻咽上皮且只在其中表达和/或在鼻咽癌HNE1中表达降低，并在鼻咽癌病人中存在不同程度的缺失.序列分析结果表明这些差异性表达的基因为具有相当抑癌基因功能的已知基因和可能与鼻咽癌相关的抑癌基因的新基因.从而说明cDNA RDA是一种高效、敏感、假阳性低的克隆抑癌基因的有效方法.

Screening the Candidates of Tumor Suppressor Genes in Human Carcinoma by cDNA Representational Difference Analysis

Using cDNA from normal human primary cultures of nasopharyngeal epithelial cells as tester amplicon and cDNA from nasopharyngeal carcinoma(NPC) cell line HNE1 as driver amplicon, four differential cDNA fragments were isolated by cDNA representational difference analysis(RDA).While these differential cDNA fragments were hybridized to amplicons by DNA blot, it was found that they really came from the tester amplicon.In addition,RNA blot also revealed that the differentially expressed fragments were not expressed or down regulated in the NPC HNE1 cells;and sequence analysis also indicated that those differentially expressed fragments may include tumor suppressor gene relative to NPC. Therefore,it shows that the cDNA RDA is an effective,sentitive and specific method for screening the candidates of tumor supressor gene.