肺癌相关基因HLCDG1的克隆和表达分析  

Cloning and Expression Analysis of Lung Carcinoma Related Gene HLCDG1

作　　者：谢海龙[1] 陈主初[1] 何春梅[1] 李友军[1] 邹飞雁[1] 关勇军[1] 

机构地区：[1]中南大学湘雅医学院肿瘤研究所细胞生物研究室,湖南长沙410078

出　　处：《癌症》2003年第10期1014-1017,共4页Chinese Journal of Cancer

基　　金：国家自然科学基金(No.30000074)

摘　　要：背景与目的:比较基因组杂交和微卫星多态性位点全基因组扫描结果显示肺癌患者在染色体3p、5q、6q、9p、10q、11p、13q、17p、19p等区域存在高频率的杂合性缺失,提示可能有多个未知的易感基因或抑癌基因与肺癌的发生、发展相关。本研究选用在mRNA差异显示研究中获得的在肺癌组织中表达下调且代表新基因的表达序列标签(expressedsequencetag,EST)LXDD1为进一步研究对象,克隆其全长cDNA。方法:采用Northern杂交验证LXDD1在肺癌中的表达差异,并用MTN(MultipleTissueNorthernBlotsTM)膜检测其在正常组织中的表达及其所代表基因的转录本大小;克隆其全长cDNA,并利用生物信息学对该序列进行初步分析;用差异RT-PCR检测新基因在肺癌组织、配对癌旁非癌组织、肺癌细胞系和其它肿瘤细胞系中的表达。结果:成功地克隆了一个在肺癌中表达下调的新基因HLCDG1(humanlungcarcinomadeletedgene1),GenBank登录号为AF447582,全长cDNA为3.113kb,预测其开放阅读框编码一个含166个氨基酸的跨膜蛋白质,通过电子-聚合酶链反应(electric-polymerasechainreaction,e-PCR)将该基因定位于5q33。结论:HLCDG1基因是一个在肺癌中表达下调的新基因,这提示HLCDG1可能与肺癌的发生、发展相关。BACKGROUND &OBJECTIVE:High frequent loss of hetero zygosity (LOH) of 3p, 5q, 6q, 9p, 10q, 11p, 13q, 17p, and 19p in lung carcinoma was detected by comparative genomic hybridization (CGH) and genomic wide scan for analysis of genetic alteration with microsatellite allelotying. It was indicated that there might be some other unknown tumor susceptible genes or suppressor genes likely to be involved in lung carcinoma development and progression. The aim of this study was to clone the full length cDNA of LXDD1,an expressed sequence tag(EST) isolated by mRNA differential display, which is significantly down regulated in lung carcinoma and represents a novel gene. METHODS: Differential expression of LXDD1 in lung carcinoma was confirmed by Northern blot analysis, the expression of the LXDD1 in human normal tissues and the size of the transcription of the LXDD1 representative gene were also determined using MTN ( Multiple Tissues Northern BlotsTM ). The putative full length cDNA of the EST representative gene was cloned and analyzed by bioinformatics. In addition, differential reverse transcription polymerase chain reaction (RT PCR) was used to detect the expression of the novel gene in various cancer cell lines, primary lung carcinomas, and matched normal lung tissues. RESULTS: The full length cDNA with no homology to any reported genes in the database of GenBank was successfully cloned and named HLCDG1 (Human lung carcinoma deleted gene 1, GenBank accession number AF447582). A transmembrane protein with 166 amino acids was deduced to come from the open reading frame of the 3113 bp full length cDNA, HLCDG1 gene was confirmed to be located at chromosome band 5q33 by alignment of electric polymerase chain reaction (e PCR). CONCLUSION: HLCDG1 is a novel gene down regulated in lung carcinoma, which may be involved in the development of lung carcinoma.

关 键 词：肺癌 HLCDGl 基因克隆 基因表达 基因组杂交 微卫星 基因多态性 

分 类 号：R734.2[医药卫生—肿瘤] R394[医药卫生—临床医学]