Livin shRNA真核表达载体构建及其对HepG2细胞化疗敏感性的影响-论文.pdf

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1、5I卷第l2期LLI东大学学报(医学版)201312J1]VO1．51NO．12JOURNALOFSHANDONGUNIVERSITY(HEALTHSCIENCES)Dec．2Ol3文章编号：1671—7554(2013)12—0020—05DOI：10．6040／j．issn．1671-7554．0．2013．011LivinshRNA真核表达载体构建及其对HepG2细胞化疗敏感性的影响徐威，常宏，翟云鹏(山东大学附属省立医院普外科，济南250021)摘要：目的构建针对Livin基因的短发夹RNA(shRNA)真核表达载体，探讨其对肝肿瘤HepG2细胞化疗敏感性的影响。

2、方法设计并构建针对Livin基因的shRNA真核表达载体pSD11一u6／Neo／GFP／Livin，脂质体法转染肝肿瘤HepG2细胞。荧光定量PCR、Westernblotting检测细胞LivinmRNA和蛋白的相对表达水平。顺铂(2．0mg／L)处理后，MTT法检测细胞生长抑制率。结果测序分析证实针对Livin基因的shRNA真核表达载体构建成功，转染肝肿瘤HepG2细胞后可降低LivinmRNA和蛋白的相对表达量(P<0．05)，顺铂对转染Livin表达载体的HepG2细胞抑制率明显高于未转染细胞(P<0．05)。结论成功构建LivinshRNA真核表达载体，且有

3、效抑制HepG2细胞中Livin基因的表达，增强肝肿瘤细胞对化疗药物的敏感性。关键词：Livin基因；RNA干扰；肝细胞肝肿瘤中图分类号：R735．7文献标志码：AConstructionofLivinshRNAeukaryoticexpressionvectoranditsinfluenceonchemotherapeuticsensitivityofHepG2cellsXUWei，CHANGHong，ZHAIYun·peng(DepartmentofGeneralSurgery，ShandongProvincialHospitalAfiliatedtoShandong

4、University，Jinan250021，China)Abstract：ObjectiveToconstructashorthairpinRNA(shRNA)eukaryoticexpressionvectortargetingtotheLivingene，andexploreitsinfluenceonchemotherapeuticsensitivityofHepG2cells．MethodsTheLivinshRNAeukaryoticexpressionvectorspSDI1一U6／Neo／GFP／livinweredesigned，constructeda

5、ndtransfectedstablyintoHepG2cells．TheLivinmRNAandproteinrelativeexpressionweredetectedbyfluorescencequantitativePCRandWesternblottingMTT．wasusedtoevaluatetheinhibitionrateofHepG2cellsafterexposuretocisplatin(2．0mg／L)．ResultsTheDNAse—quencingconfirmedthattheLivinshRNAeukaryoticexpressionve

6、ctorsweresuccessfullyconstructed．TransfectionofthevectorscouldsignificantlydecreasetheLivinmRNAandproteinrelativeexpressionofHepG2cells(P<0．05)．TheinhibitionrateofcisplatinintransfectedcellswereobviouslyhigherthanthoseintheuntransfectedcellsfP<0．05)．ConclusionTheshRNAeukaryoticexpressionv

7、ectorstargetingtotheLivingeneareconstructedsuccessfully．TransfectionofthevectorsCanefectivelydecreasetheexpressionofLivingeneinHepG2cells，andincreasethesensi—tivityofHepG2cellstochemotherapy．Keywords：Livingene；RNAinterference；HehapocellularneoplasmLivin是凋亡抑制蛋白(inhib