鼻咽癌中差异表达基因的分析和克隆  

Analysis and Molecular Cloning of Differentially Expressing Genes in Nasopharyngeal Carcinoma

作　　者：余鹰[1] 张必成 谢奕[1] 曹利[1] 周鸣[1] 湛凤凰[1] 李桂源[1] 

机构地区：[1]湖南医科大学肿瘤研究所,长沙410078

出　　处：《生物化学与生物物理学报》2000年第4期327-332,共6页

基　　金：国家高科技"863"计划资助项目!No.10 2 10 0 1 0 5 ;国家"973"重点项目资助;国家自然科学基金资助项目!No .3 970 0 15 8&&

摘　　要：为了验证通过cDNA代表性差异分析法 (cDNArepresentationaldifferenceanalysis ,cDNARDA)分离的cDNA序列在鼻咽癌活检组织中的差异表达 ,并进一步克隆鼻咽癌差异表达基因 ,采用RT PCR方法并结合Nor thern杂交确定其转录本的大小 ,进而充分利用生物信息学资源对有差异表达cDNA全长进行克隆 ,并对其基因产物进行了结构和功能的预测。结果显示 ,AF0 915 2 1、AF0 915 2 0、AF15 2 6 0 5、AF0 915 17等cDNA序列在正常鼻咽上皮与鼻咽癌组织中存在表达差异 ;AF0 915 2 1和AF0 915 17基因均有 2个转录本 ,大小分别为 1.5、2 .3和1.1、1.4kb ;AF0 915 2 0和AF15 2 6 0 5基因转录本的大小分别为 1.6和 2 .2kb。结合生物信息学直接测序 ,克隆了包含AF0 915 17EST的基因 ,命名为NAG11,GenBank收录号为AF170 30 7,其编码 88氨基酸组成的跨膜蛋白 ,含有 3个蛋白ATP结合区和 2个蛋白激酶C磷酸化位点和 2个N 肉豆蔻酸化位点。以上结果进一步证明鼻咽癌是一种多基因性疾病 ;AF0 915 2 1、AF0 915 2 0、AF15 2 6 0 5、AF0 915 17可能代表某些鼻咽癌易感 /抑瘤基因的候选者 ;NAG11基因的下调参与了鼻咽癌的发生、发展 ;NAG11基因产物可能与ATP的跨膜转运有关。In order to further examine expression of cDNA fragments isolated by cDNA representational difference analysis(cDNA RDA) in nasopharyngeal carcinoma(NPC) biopsies and to clone those deregulated genes associated with NPC, RT PCR and Northern blot were used to identify the differentially expressed cDNA fragments in NPC biopsies and confirm the transcript length of those genes, then a full length cDNA sequences was cloned and its product was analyzed by bioinformatics. The results showed that AF 091521, AF 091520, AF 152605 and AF 091517 cDNA sequences had distinct expression difference between primary cultural normal nasopharyngeal epithelial cell and NPC biopsies, and AF 091521, AF 091517 genes all had two transcripts whose sizes were 1.5, 2.3 and 1.1, 1.4 kb respectively, while AF 091520 and AF 152605 gene expressed one trans cript only, respectively, whose sizes were 1.6 and 2.2 kb. An AF 091517 EST gene, named as NAG 11, (GenBank accession number: AF170307) was isolated by sequencing one EST clone, which encoded a transmembrane protein of 88 amino acid including three protein ATP binding regions, two protein kinase C phosphorylation sites and two N myristolation sites. So it is further demonstrated that NPC is a disease with multiple gene alterations; NAG 11 gene is a candidate of putative tumor suppressor genes associated with NPC, whose down expression may be involved in the development of NPC; and NAG 11 gene product may play a role in the transmembrane transport of ATP.

关 键 词：基因表达 抑瘤基因 鼻咽癌 CDNA RDA 基因克隆 

分 类 号：Q785[生物学—分子生物学]