【摘要】 目的　克隆野生型 p5 3 (wtp5 3 )影响人大细胞肺癌 80 1 D细胞系顺铂敏感性相关基因片段。 方法　采用转基因方法补偿 80 1 D细胞p5 3功能 ,分别提取经IC50 浓度顺铂刺激的转染wtp5 3细胞、空载细胞和未经顺铂作用的转wtp5 3细胞RNA ,用银染mRNA差异显示方法分离差异表达基因 ,反转录点杂交和巢式PCR进行确证。阳性片段进行克隆及序列分析。结果　获得 6条阳性片段 ,2条含有开放阅读框架(ORF) ,其中 1条片段部分与核糖核苷还原酶α链有 5 6%同源性 ,另一片段D1含有一个长ORF ,与已知cDNA序列同源率低。结论　p5 3对 80 1 D细胞顺铂敏感性的影响有明显的基因水平改变 ,可能诱导了相关基因表达。更多还原

【Abstract】 Objective To isolate and clone the cisplatin genes in 801 D cell line, a kind of lung cancer cell line, with the emphasis of the objective genes regulated by wild type p53 (wtp53). Methods Total RNA was extracted from transfected 801 D wtp53, 801 D vector cells which were both treated by cisplatin and 801 D wtp53 cells. Using mRNA differential display, the DNA bands on gel were displayed by silver stain method. The DNA bands obtained from differential display were recovered and reamplified by PCR. The isolated genes were further proved by reverse Northern dot blot and were cloned to pGEMT easy vector. Results Six positive genes were identified and cloned. Out of them, 2 related fragments were found to have an open reading frame. One was partly homologous to ribonucleoside diphosphate reductase A, and the other was no homologous to the known genes. Conclusion There are obvious differences in gene expression in 801 D wtp53 after induced by cisplatin than two other controls. It is possible for p53 to regulate the sensitization of lung cancer cells to cisplatin through its downstream target genes.更多还原