摘要
目的本研究试图利用前期工作中诱导建立的小细胞肺癌多药耐药细胞系构建小细胞肺癌多药耐药细胞差异表达消减cDNA文库,为进一步功能研究奠定基础。方法 (1)以小细胞肺癌多药耐药细胞H446/CDDP cDNA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。方法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%)。方法本研究表明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。
Objective To construct the subtracted cDNA library of differentially expressed genes of small-cell lung cancer multi-drug resistance （ SCLC MDR） cell by using a cell line created previously. Methods （ 1 ） The tester was SCLC MDR cell H446/CDDP cDNA and the driver SCLC cell H446 cDNA. Suppression subtractive hybridization （SSH） and T/A cloning technology were applied to construct the subtracted cDNA library of differentially expressed genes of SCLC MDR cell in this research. （2） The dot blot hybridization and sequencing and homology analysis were used to obtain differentially expressed cDNA fragments. Results （1） We successfully constructed the subtracted cDNA library of differentially expressed genes of SCLC MDR cell H446/CDDP and obtained 21 differentially expressed cDNA fgragments of cell H446/CDDP. （2） Sequencing and homology analysis showed that the 21 fragments respectively represented 6 known genes （96% ～ 100% homology）. Conclusion SSH is an effective method for screening new functional genes. The obtained 6 differentially expressed genes might participate in the formation of the MDR in H446/CDDP. Further studies are needed to determine how the six genes function in SCLC MDR mechanism.
出处
《中华肺部疾病杂志（电子版）》
CAS
2007年第1期32-36,共5页
Chinese Journal of Lung Diseases(Electronic Edition)
基金
国家自然科学基金资助项目(30200119)
关键词
小细胞肺癌
多药耐药
抑制消减杂交技术
Small cell lung cancer
Multi-drug resistance
Suppression subtractive hybridization
