ISOLATION OF POTENTIAL ONCOGENES FROM TELEOST TUMORS

虽然肿瘤发生的详细机制尚不清楚，但越来越多的研究表明， 有证据表明，细胞癌基因的遗传改变， 部分，负责细胞的肿瘤转化。一些研究 在啮齿类动物模型中， 致癌物暴露。重复性检测特异性 动物模型系统中的转化基因强烈表明， 基因在某些肿瘤的发展中起重要作用。数据 从我们初步的转染实验中， 致癌物诱导的青鳉肿瘤支持这一假设。我们的数据表明 一个未知的基因可能在DEN诱导的细胞中被激活， 胆管癌它似乎与任何已知的 基于Southern印迹杂交的癌基因序列。平行 不添加pSV 2neo的二级和三级转染 揭示了在一些转染的细胞中存在质粒序列， 与细胞转化高度相关。这表明pSV 2neo可能 整合到宿主基因组中，并与宿主基因组紧密相连。 推定的鱼类癌基因，并可用作标记来鉴定该基因。 这种明显新的癌基因的鉴定和特征 正在进行中。从以下的EcoRI部分序列制备文库： 将三级转染子DNA连接到EMBL 4载体中。图书馆是 用pSV 2neo筛选，分离出12个阳性克隆，目前正在进行 表征了还开始研究开发转基因 通过引进E. colilacZ基因在 小鼠Mc-I金属硫蛋白启动子。基因表达将被诱导， 将鱼暴露于重金属中，并通过比色测定进行分析 使用靛蓝底物X-gal。这一结构也是 引入鱼类细胞系，形成体外毒性的基础 测定。初步测定了对几种有毒金属的含量 细胞系正在开发中。目前正在对这些转染子进行分析。

While the detailed mechanisms of tumorigenesis are unknown, increasing evidence suggests that genetic alterations of cellular oncogenes are, in part, responsible for the neoplastic transformation of cells. Some studies in rodent models have implicated the direct chemical activation of oncogenes by carcinogen exposure. The reproducible detection of specific transforming genes in animal model systems strongly suggests that these genes have a significant role in the development of certain tumors. Data from our preliminary transfection experiments with DNA from carcinogen-induced medaka tumors support this hypothesis. Our data suggest that an unknown gene may be activated in the DEN-induced cholangiocarcinoma. It does not appear to have homology to any known oncogene sequence based on hybridization of Southern blots. Parallel secondary and tertiary transfections without the addition of pSV2neo revealed the presence of plasmid sequences in some transfected cells, highly correlated with cell transformation. This suggested that pSV2neo may be integrated into the host genome and tightly physically linked to the putative fish oncogene, and may be used as a marker to identify this gene. The identification and characterization of this apparently novel oncogene is now in progress. A library was prepared from EcoRI partial digests of tertiary transfectant DNA ligated into an EMBL 4 vector. The library was screened with pSV2neo; 12 positive clones were isolated and are now being characterized. Studies have also been initiated to develop transgenic medaka by the introduction of the E. coli lacZ gene under the control of the mouse Mc-I metallothionein promoter. Gene expression will be induced by exposure of the fish to heavy metals and analyzed by a colorimetric assay using the indigogenic substrate, X-gal. This construct has also been introduced into fish cell lines to form the basis of in vitro toxicity assays. Preliminary determinations of levels of metals toxic to several cell lines are in progress. These transfectants are now being analyzed.