资源描述：
《基因枪转导突变特异性k-ras sirna对胰腺癌细胞生长的抑制作用》由会员上传分享，免费在线阅读，更多相关内容在教育资源-天天文库。
1、基因枪转导突变特异性K-rassiRNA对胰腺癌细胞生长的抑制作用?28?中华实验处科蠢2006年1月第23卷第1期ChinJExpSu喁January2006.Vol23.No.1基因枪转导突变特异性K—rassiRNA对胰腺癌细胞生长的抑制作用张峰刘晔刘三光谢绍建李冬斌李荣琴蔡建辉【摘要】目的探讨基因枪转导突变特异性K.rassiRNA对胰腺癌细胞生长的抑制作用.方法根据胰腺癌细胞系突变特征,设计突变特异性K,rassiRNA;应用基因枪将siRNA转导入胰腺癌细胞系,提取总RNA和胞浆蛋白;应用逆转录一聚合酶链反应(RT-PCR)和免疫印迹法(We
2、sternblot)方法.检测基因枪转导突变特异性K,rassiRNA对突变型K.ra8基因表达的干扰作用;行细胞爬片K.ra$p21蛋白免疫组织化学染色,探讨siRNA对K.ra$p21蛋白表达的抑制作用;采用CCK.8(ceIlcounringkit,8)活细胞计数法,绘制转基因前后细胞生长曲线,观察siRNA的抑瘤效果.结果RT.PCR结果显示,K,ra8突变特异性siRNA,能有效抑制K.ra8基因表达(P<0.05);Westemblot和免疫组织化学结果表明,K,ra$p21蛋白的表达明显降低(P<0.05);细胞生长曲线显示,转
3、导siRNA组较对照组细胞生长明显受抑制(P<005).结论基因枪可以有效地对小片段双链siRNA进行细胞内转导;突变特异性K—ra8siRNA,可显着下调胰腺癌细胞系突变型K.ra8mRNA及K.ra8p21蛋白表达.并能有效抑制肿瘤细胞的增殖.【关键词】胰腺癌;RNA干扰;基因枪;基因治疗ExperimentalstudyofK-rasgeneinterferenceandtumorgrowthinhibitionbygeneguntransductionofmutationspecificK-rassiRNAZH4NGFeng,L儿,L儿Sa
4、n?guang,eta1.DepartmentofSurgery,SecondAffiliatedHospital,HebeiMedicalUniversity,Shijiazhuang050000,ChinaCorrespondingauthor:c氏lJan.hui【Abstract】ObjectiveToinvestigatetheinhibitoryeffectofthegrowthofpancreaticcarcinomacelllinesafterK—ra$sirNAtransductionbygenegun.MethodsUndertheo
5、ptimumConditionsoftrans.duction,pre-designedK.rassiRNAwastransducedintopancreaticcarcinomaceillines.Afterculturefor24h,RT-PCRandWesternblotanalysiswerecarriedouttoanalyzetheinhibitoryeffectsoftumorcellgrowthtransduetedwithK?rassiRNAbygenegun.Immunohistochemistrywasperformedtodete
6、cttheexpressionofK—rasp21inpancreaticcarcinomacelllines.TheceilgrowthactivitieswereestimatedbyCCK,8cel1Droliferationassay.ResultsRT,PCRrevealedthattheceilstreatedwithsiRNAhadlowertranscriptionofK,rasmRNAthanthecontro1.Westernblotsandimmunohistochemistryindicatedade.creasedexpress
7、ion,asComparedwiththecontrols,ofK—rasproteininthecellstransductedbysiRNA.Mu.tationspecificK—ra8antisenseoligonucleatidesexhibitsananti—proliferateeffectincellproliferationassay.ConclusionK-rassiRNAcansignificantlydown—regulatethetranscriptionandexpressionofKra$inmutationpositivep
8、ancreaticcarcinomaceillinesaftereffectiv
