人肝癌细胞差异表达基因纤维蛋白原γ-多肽的克隆与鉴定  

Cloning and Identification of Fibrinogen Gamma Polypeptide (FGG) Gene Differentially Expressed in Human Hepatocellular Carcinoma

作　　者：范秉琳[1] 朱武凌[1] 邹国林[2] 雒国胜[1] 许春雷[1] 赵卫星[1] 

机构地区：[1]新乡医学院基础医学部,河南新乡453003 [2]武汉大学生命科学学院生物化学教研室,湖北武汉430072

出　　处：《癌症》2004年第3期249-253,共5页Chinese Journal of Cancer

基　　金：河南省高校青年骨干教师资助项目(No.2001225158)~~

摘　　要：背景与目的:肝细胞癌(肝癌)的发生、发展与基因的异常表达有关,但详细机制还不清楚,原因在于目前所知的遗传学信息尚欠充分;本研究旨在探索新的人肝癌细胞差异表达基因。方法:应用抑制性消减杂交技术获得肝癌细胞消减cDNA,以T/A方法进行克隆,通过DNA测序分析、Northern印迹杂交、cDNA末端快速扩增(rapidamplificationofcDNAend,RACE)和RT-PCR等方法加以鉴定。结果:在被克隆的消减cDNA之中,一个长度为787个核苷酸的差异表达cDNA片段经Northern印迹分析,发现其在两种人肝癌细胞株SMMC-7721和HepG2中的表达均明显高于正常肝细胞;经过RACE后获得一个长度为1597bp、具有3'端polyA结构的基因序列,它含有完整的开放阅读框,编码437个氨基酸,经同源性分析证实该基因为编码人纤维蛋白原γ-多肽(fibrinogengammapolypeptide,FGG)的基因;RT-PCR分析证明癌组织中FGGmRNA表达水平较癌旁非癌组织明显增加,尤其是在肝癌转移灶中。结论:SMMC-7721和HepG2细胞中发现FGG高表达,FGG基因的上调表达是肝癌病理过程中一个重要分子事件。BACKGROUND & OBJECTIVE: Abnormal expression of genes is related t o development and progression of hepatocellular carcinoma (HCC); however, the de tailed mechanism is unclear yet because the known genetic information is not suf ficient at present. This study was to explore cloning and identification of fibr inogen gamma polypeptide (FGG) gene differentially expressed in human hepatocell ular carcinoma. METHODS: The suppression subtractive hybridization was used to o btain subtracted cDNA products of HCC, then the products were cloned by T/A meth od. The differential expression of gene in HCC was identified by DNA sequencing analysis, Northern blot analysis, rapid amplification of cDNA end (RACE) , and r everse transcription polymerase chain reaction (RT-PCR). RESULTS: Firstly, a cD NA fragment of 787 nucleotides was screened from the subtracted cDNA clones, and it was further discovered that the expression of the cDNA fragment was higher s ignificantly in human hepatocellular carcinoma cell strains of SMMC-7721 and He pG2 than in normal hepatocytes by Northern blot analysis. The RACE was carried o ut and the gene of 1 597 bp containing polyA in 3′end was obtained, which has a n entire open reading frame encoding 437 amino acids. Homology analysis showed t hat this was a gene encoding human FGG. RT-PCR analysis of FGG showed that the amplification of cancerous tissues, especially in metastasis of HCC, was raised as compared to that of adjacent non-cancerous tissues. CONCLUSION: Overexpressi on of FGG was discovered in SMMC-7721 and HepG2 cells. The up-regulation of FG G may be associated with the pathogenesis of HCC.

关 键 词：肝癌细胞 基因纤维蛋白原 γ-多肽 克隆 基因表达 肿瘤 

分 类 号：R735.7[医药卫生—肿瘤]