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目的 :利用mRNA差异显示技术研究正常卵巢上皮和卵巢上皮癌之间的基因表达差异状况 ,筛选和克隆卵巢上皮癌相关基因。方法 :通过DD PCR分析正常卵巢上皮和卵巢上皮癌总RNA将差异片段分离并显示出来 ,将其回收 ,利用基因克隆技术将其筛选和克隆出来作为探针进行斑点杂交鉴定、DNA序列测定、通过国际互联网络检索GenBank进行同源性分析。结果 :共筛选克隆鉴定 6个差异显示片段。其中 4个为未知基因片段 ,已收录入GenBank ,GenBank登录号为AF136 413～AF136 417。两个为已知基因片段 ,其中一个克隆 3AOSKN3与GenBank中染色体 17hCIT克隆 98%同源。另外一个克隆 3AON2 8与GenBank中人类NADH :辅酶Q氧化还原酶有 99%的同源性 ,研究表明该基因与肿瘤相关。结论 :上述结果证明mRNA差异显示方法和杂交分析结合已成为一种敏感、高效、快速的差异表达基因的克隆技术
OBJECTIVE: To study the difference of gene expression between normal epithelial ovarian epithelial carcinoma and ovarian epithelial carcinoma by mRNA differential display technique, and to screen and clone the gene of ovarian epithelial carcinoma. Methods: The total RNA of normal ovarian epithelial carcinoma and ovarian epithelial carcinoma was separated and displayed by DD PCR. The differential fragments were recovered and recovered. The cDNA fragments were screened and cloned by using gene cloning technique to identify dot blot hybridization and DNA sequencing. The Internet searches GenBank for homology analysis. Results: A total of 6 differential display fragments were identified by screening clones. Four of them are unknown gene fragments and have been included in GenBank. The GenBank accession numbers are AF136 413 ~ AF136 417. Two are known gene fragments. One of the clones, 3AOSKN3, is 98% homologous to the chromosome 17hCIT clone in GenBank. Another clone, 3AON2 8, has 99% homology with the human NADH: coenzyme Q oxidoreductase in GenBank. Studies have shown that the gene is tumor-associated. CONCLUSIONS: The above results demonstrate that mRNA differential display and hybridization assays have become a sensitive, efficient and rapid cloning technique for differentially expressed genes