Cloning of genes expressing in the early stage of rat bladder carcinogenesis induced by BBN

BBN诱导大鼠膀胱癌早期表达基因的克隆

• 批准号: 06670224
• 负责人: ONO Takahiko
• 金额: $ 1.41万
• 依托单位: KYOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06670224/
• 关键词: Urethelial cancer; Tumor suppressor; Carcinogenesis; Cloning; PCR; Chemical carcinogenesis; ラット発癌; BBN; 遺伝子クローニング; LOH

We have been studying inactivation of tumor suppressor genes in human urothelial cancer cells. Along with analyzes using human tumor tissues, experimental models should provide data in the early stage of carcinogenesis or in precancerous lesions. A recently developed technique of differential RT-PCR allows us to detect expression of genes highly expressed in such primary lesions. It would also be possible to identify human homologues from clones isolated by the differential RT-PCR method. Total cytoplasmic RNA was isolated from hyperplastic lesions, papilloma, and carcinoma in situ of the rat bladder treated with BBN for six months. Differential display was performed with several sets of random primers and oligo dTs using a normal bladder mucosa as a control. Forty-two bands were chosen because they were specificly found or their intensity was increased. Confirmation of specific expression in each lesions was performed using Northern blot or quantitative RT-PCR assay. Due to a limited amount of RNA from the primary lesions, quantitative RT-PCR was effective for such evaluation. Further characterization of the isolated clones are ongoing.

我们一直在研究人尿路上皮癌细胞中抑癌基因的失活。沿着使用人肿瘤组织的分析，实验模型应提供癌变早期或癌前病变的数据。最近开发的差异RT-PCR技术使我们能够检测在这种原发性病变中高度表达的基因的表达。也有可能从通过差异RT-PCR方法分离的克隆中鉴定人类同源物。从用BBN治疗6个月的大鼠膀胱增生性病变、乳头状瘤和原位癌中分离总细胞质RNA。用几组随机引物和寡脱氧核糖核酸进行差异显示，正常膀胱粘膜作为对照。选择42条谱带，因为它们是特异性发现的或它们的强度增加。使用北方印迹或定量RT-PCR测定来确认每个病变中的特异性表达。由于来自原发性病变的RNA量有限，定量RT-PCR对于此类评估是有效的。分离克隆的进一步表征正在进行中。

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