论文部分内容阅读
目的 探讨人胰腺淀粉酶基因2(Amy-2)启动子在人胰腺组织特异性并验证其在胰腺细胞内的活性.方法 运用荧光素酶报告载体构建胰腺组织特异性启动子Amy-2质粒,将其转染人胰腺癌细胞系SWl990、PANc-1、正常人胰腺导管上皮细胞(HPDE)、乳腺癌(MCF-7)、肺癌(A549),验证其在胰腺细胞内的活性和特异性.利用分子生物学技术,结合PBS185载体构建载体pAmy2-Cre;在运用基因转染方法 将pAmy2-Cre的转染人胰腺癌细胞株Pane-1、BxPC-3,通过PCR、Western blot方法 观察pAmy-2的组织特异性.结果 成功构建pAmy-2-1uc和pAmy-2-Cre载体,证明了Amy-2的启动子在相应细胞内的转录活性;Amy-2的启动子能很好的驱动Cre蛋白表达.结论 Amy-2有较好的胰腺组织特异性和转录.“,”Objective To examine the efficacy of gene under the control of pancreatic-tissue-specific promoter of human amylase-2 gene (Amy-2). Methods Transcriptional activities of Amy-2 promoter sequences were analyzed using a luciferase reporter gene on human pancreatic cancer cell lines SW1990 and PANC-1, human pancreatic duct epithelial cell (HPDE), human breast cancer cell line MCF, human lung cancer cell line A549 cell lines. The recombinant pAmy-2-Cre vector was constructed with PBS185 vector. The efficacy of pAmy-2-Cre on the transiently trancfected Panc-1 and BxPC-3 human pancreatic cancer cell lines. Expression of Cre gene of transiently trancfected cells was measured by Western blot assay in vitro. Results The Amy-2 promoter had high transcriptional activity in transfected tumor cell-lines and pancreas-derived cells respectively. Expression of Cre gene with pAmy-2-Cre vector was proved to be tissue-specific promoter Amy-2 regulated. Conclusion The promoter Amy-2 has high transcriptional activities and tissue-specific activity in pancreatic cancer cells.