食管癌低表达cDNA片段C6-2A的克隆及表达分析

Cloning and expression analyses of down regulated cDNA C6 2A in human esophageal cancer

目的 克隆人食管癌发生相关的基因。方法 用ｍ Ｒ Ｎ Ａ 差异显示技术分离、克隆食管癌组织中不表达或低表达的ｃ Ｄ Ｎ Ａ 片段，再通过 Ｎｏｒｔｈｅｒｎ ｂｌｏｔ 、ｄｏｔ ｂｌｏｔ 和 Ｒ Ｔ Ｐ Ｃ Ｒ 证实。结果 获得２８０ｂｐ 的ｃ Ｄ Ｎ Ａ 片段，命名为 Ｃ６２ Ａ。与 Ｇｅｎ Ｂａｎｋ 基因数据库比较，未发现 Ｃ６２ Ａ 与任何已知基因有同源性。查询 Ｅ Ｓ Ｔ 数据库，发现 Ｃ６２ Ａ 与ｎｅ２７ｂ０３ ，ｓ１ Ｎ Ｃ Ｉ Ｃ Ｇ Ａ Ｐ Ｃ０３ 人ｃ Ｄ Ｎ Ａ 克隆８９８５４１ 及人卵巢肿瘤ｃ Ｄ Ｎ Ａ 克隆７５５１９６ 等高度同源。 Ｎｏｒｔｈｅｒｎ ｂｌｏｔ 结果显示６／６ 例食管癌组织表达丧失或低表达，ｄｏｔ ｂｌｏｔ 分析表明７／８例食管癌组织低表达， Ｒ Ｔ Ｐ Ｃ Ｒ 显示食管癌细胞系 Ｅ Ｃ１０９ 、 Ｅ Ｃ８７１２ 、 Ｅ Ｃ９７０６ 和肺腺癌细胞系 Ｇ Ｌ Ｃ８２ 极弱表达，１７／２０ 例食管癌组织表达丧失或低表达，胎儿食管、皮肤、大脑、胎盘较高表达，胎儿胃、肝较弱表达，胎儿心、小肠、肾不表达。结论 在食管癌细胞系和食管癌组织高频率的不表达或低表达提示， Ｃ６２ Ａ 很可能与食管癌的发生、发展有关。

Objective To clone genes associated with the genesis of human esophageal cancer.Methods Identifying missing or low expressing cDNAs in human esophageal cancer tissues by mRNA differential display and examining its mRNA expression in 4 human cancer cell lines, 9 fetal tissues and other matched esophageal cancer tissues by Northern blot, dot blot and RT PCR.Results One cDNA fragement named C6 2A, was cloned and sequenced. There was no identical sequence with C6 2A in BLASTN database; but in querying Genbank EST, the authors found that C6 2A was identical with ne27b03.s1NCI CGAP C03 humans sapiens cDNA clone IMAGE:898541 3′ and zv30g07.r1 Soares ovary tumor NbHOT homo sapiens cDNA clone 755196.6/6 esophageal cancer tissues in Northern blot and 7/8 in dot blot did not or slightly express C6 2A. RT PCR analysis showed that C6 2A was expressed much lower in 17/20 espohageal cancer tissues than adjacent microscopically normal mucosa, highly expressed in fetal esophageal mucosa, skin, cerebrum,placenta;moderately expressed in fetal stomach and liver, but not detected in fetal heart, small intestine and kidney. Conclusion The high frequency of deletion of decreased expression of C6 2A in esophageal cell lines and human esophageal cancer tissues suggested that C6 2A might be involved in the carcinogenesis of esophagus.