摘要
目的:用脂质体法建立nucleostemin(NS)基因特异性siRNA阳性细胞克隆,为深入研究NS基因在胃癌细胞增殖调控中的作用和机制提供理想的生物学模型。方法:试剂盒法将真核表达载体PCDNA4/C-NS-silencer质粒大量扩增,然后以限制性切酶PVUI酶切线性化处理,用LipofectamineTM2000 Reagent将PCDNA4/C-NS-silencer及其对照空载体转染胃癌SGC-7901细胞,经Zeocin抗生素压力筛选后用PCR方法鉴定细胞克隆外源基因整合阳性。结果:经Zeocin抗生素压力筛选后两组细胞均收获多个细胞克隆;PCR结果表明两组细胞克隆外源基因整合阳性。结论:成功建立表达NS-siRNA的胃癌SGC-7901细胞克隆。
Objective： To screen NS gene specific positive cell clones, making a model for the research on the function of NS gene in human Gastric Carcinoma SGC -7901 cells proliferous regulation. Methods：After the eukaryotic NS -siRNA expression vector was digested by PVUI restriction enzyme , transfected it into human Gastric Carcinoma SGC -7901 cells using LIPOFECTAMINETM2000 Reagent, and cell clones were screened with Zeocin. We isolated genomic DNA from these cell clones and examined wether integration with exogenous gene by PCR. Results： Cell clones expressing NS - siRNA were acquired after screening with Zeocin, PCR result showed that cell clones integration with exogenous gene positive. Conclusion： Constructed NS - siRNA of human Gastric Carcinoma SGC - 7901 cells clones.
出处
《牡丹江医学院学报》
2009年第4期5-8,共4页
Journal of Mudanjiang Medical University
基金
黑龙江省普通高等学校青年学术骨干支持计划项目(1153G047)
黑龙江省卫生厅科研项目(2007-021)
牡丹江医学院一般科研项目(B200822)
