人小细胞肺癌多药耐药细胞差异表达基因的克隆研究

目的 构建小细胞肺癌多药耐药细胞(H446/CDDP)差异表达消减cDNA文库.方法 ①以H446/CDDPcDNA为实验方(Tester),小细胞肺癌细胞H446 cDNA为对照方(Driver),应用抑制消减杂交(SSH)技术结合T/A克隆技术构建小细胞肺癌多药耐药细胞差异表达消减cDNA文库.②通过斑点杂交筛选、测序及同源性分析获取H446/CDDP细胞差异表达cDNA片段.结果 成功构建了小细胞肺癌多药耐药细胞H446/CDDP差异表达消减cDNA文库,获得21个H446/CDDP细胞差异表达cDNA片段,经测序及同源性分析表明它们分别代表6个已知基因(同源性为96%～100%).结论 SSH技术是筛选新的功能基因的有效方法;获取的6个差异表达基因可能参与了小细胞肺癌多药耐药细胞H446/CDDP的耐药形成,进一步的功能研究有利于了解它们在小细胞肺癌多药耐药机制中所发挥的作用.

Clone of differentially expressed genes of human small-cell lung cancer multi-drug resistance cell line

Objective To build the subtracted cDNA library of differentially expressed genes of small cell lung cancer(SCLC)multidrug resistance(MDR)cells.Methods The tester is SCLC MDR cell H446/CDDP cDNA;the driver is SCLC cell H446 cDNA;SHH and T/A cloning technology were done to build the subtracted cDNA library of differentially expressed genes of SCLC MDR cells.The dot blot hybridization,sequencing and homology analysis were used to obtain differentially expressed cDNA fragments.Results The subtracted cDNA library of differentially expressed genes of SCLC MDR cells H446/CDDP was successfully built and 21 differentially expressed cDNA fgragments of cell H446/CDDP were obtained.Sequencing and homology analysis show that the 21 fragments respectively represent 6 known genes(96%-100% homology).Conclusion Suppression subtractive hybridization(SSH)is an effective method for screening new function genes.The six genes might participate in the formation of the MDR in H446/CDDP.