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1、dsRNA阻断大鼠骨髓源性神经干细胞Hes5表达的实验研究摘要：目的 观察外源性短双链RNA(dsRNA)在转录后水平即mRNA水平降低基因表达的效率，并对其影响因素进行初步探讨。方法 将体外分离培养的大鼠骨髓基质细胞，通过由本实验室配制的特殊培养基诱导成神经干细胞，应用人工合成的dsRNA于不同浓度转染神经干细胞，Westernbloting检测dsRNA是否能阻断转录因子Hes5的表达，0.5%锥虫蓝法测定各浓度组中培养细胞活力。结果 200~300nmol/L浓度的dsRNA能特异有效地阻断Hes5的表达，而50~200nmol/L浓度的dsR
2、NA有利于干细胞存活。结论 dsRNA能在神经干细胞内启动RNA干扰作用，适宜浓度的dsRNA能在特异有效地阻断内源性基因的表达的同时，最大限度地保持细胞活力。关键词：双链RNA；Hes5；神经干细胞，骨髓源性；RNA干涉；免疫印迹Blockingwithdouble-strandedRNAoftheexpressionofHes5inratbonemarrow-derivedneuralstemcellsWANGHai-yan1；XURu-xiang1；JIANGXiao-dan1；LUOShen-qiu21InstituteofNeuromedici
3、neofPLA,ZhujiangHospital,FirstMilitaryMedicalUniversity,Guangzhou510282,China;2DepartmentofCellularBiology,FirstMilitaryMedicalUniversity,Guangzhou510515,ChinaAbstract:Objective Toexaminetheefficiencyofexogenoussmalldouble-strandedRNA(dsRNA)inknockingdownthegeneexpressionatthepos
4、t-transcriptionlevel,andinvestigatethefactorsthatmayinfluence本文由探生科技技术人员提供，如果您想了解更多关于抗体的信息，可以访问Fantibody全球抗体搜索引擎，您身边的抗体专家查找相关资料,希望对您有帮助！thetransfection.Method ThebonemarrowstromalcellsofSDratwereseparatedandculturedinvitro,followedbyinductionofthecellstoevolveintoneuralstemcellsu
5、singspecialculturemediumpreparedbyourlaboratory.SyntheticdsRNAwasthentransferredintothecellsatvariedconcentrations,andtheresultswereanalyzedbyWesternblotting.Results Theconcentrationsrangingfrom200to300nmol/LwereoptimalforspecificallyblockingtheexpressionofHes5,whereasthesuitable
6、concentrationsforthecellsurvivalwerebetween50and200nmol/L.Conclusion dsRNAiscapableoftriggeringRNAinterferenceinneuralstemcells,andatappropriateconcentration,itmayspecificallyandeffectivelyknockdownendogenousgeneexpressionwithoutsacrificingtheviabilityofthecells.Keywords:double-s
7、trandedRNA;hairyandenhancerofsplit;neuralstemcell,bonemarrow-derived;RNAinterference;Westernblotting收稿日期：2003-07-11基金项目：国家自然科学基金(3027049)；军队及广东省科技厅“十五”重大项目(01z054)ThestudyissupportedbyNationalNaturalScienceFoundationofChina(3027049),andisakeyprojectoftheTenthFive-yearPlanofthePLAa
8、ndScienceandTechnologyCommissiono
