肝癌组织差异表达基因cDNA序列的筛选与鉴定  

Screening and identification for cDNA of differentially expressed genes in human primary hepatocellular carcinoma

作　　者：李靖[1] 韩本立[2] 黄桂君[3] 钱桂生[3] 梁平[1] 杨彤翰[1] 陈杰[3] 

机构地区：[1]第三军医大学新桥医院肝胆外科,重庆400037 [2]西南医院肝胆外科 [3]西南医院呼吸内科研究所

出　　处：《中华医学遗传学杂志》2003年第1期49-52,共4页Chinese Journal of Medical Genetics

基　　金：国家自然科学基金 (30 1 70 4 2 4 ) ;重庆市卫生局医学科研基金 (0 0 - 2 0 0 1 )~~

摘　　要：目的　筛选并鉴定肝癌组织特异表达基因。方法　通过菌落原位杂交技术筛选用抑制消减杂交法构建肝癌与癌旁肝组织差异表达基因消减 c DNA文库 ,用 PCR方法进一步筛选出有插入片段的阳性克隆 ,将阳性克隆进行 DNA测序和同源性比较分析 ,用 Northern印迹方法对新的 c DNA序列进行初步鉴定。结果　从消减文库中随机挑取的 10 0个白色克隆中筛选出 13个阳性克隆 ,DNA测序获得 11个不同的 c DNA序列 ;同源性比较分析表明 ,6个 c DNA片段与已知基因高度同源 ,5个 c DNA片段为新的序列。其长度大于 30 0 bp的 3个新序列 ,Northern印迹证实它们都来源于肝癌组织。结论　用抑制消减杂交方法构建的肝癌差异表达基因消减 c DNA文库富含肝癌特异表达基因 ,经验证的 3个新的 cObjective: Screening and identification of differentially expressed genes in human primary hepatocellular carcinoma (HCC). Methods: The differentially expressed genes subtracted cDNA library of HCC constructed by suppression subtractive hybridization (SSH) technique was screened by colony in situ hybridization, then the positive clones were further screened with PCR amplification. The positive clones were sequenced and analyzed for homology in the Genbank databases with Basic Local Alignment Search Tool (BLAST). The novel cDNA sequences were analyzed by Northern blot analysis. Results: Thirteen positive clones were obtained, and 11 cDNA sequences were identified. Sequences of 11 cDNA showed that 6 cDNA were homologous with the genes published in Genbank and 5 cDNA were unknown genes. Northern blot indicated that 3 novel cDNA (> 300 bp) were only expressed in HCC. Conclusion: The subtracted cDNA library constructed by SSH technique contains differentially expressed genes of HCC. Three novel cDNA sequences might be differentially expressed genes of HCC. Further screening the library and gaining the whole gene sequence may lay a foundation for identifying differentially expressed genes in HCC.

关 键 词：肝癌组织 差异表达基因 CDNA 序列 筛选 鉴定 肝肿瘤 基因表达 CDNA文库 

分 类 号：R735.7[医药卫生—肿瘤]