资源描述：
《jagged1真核表达质粒构建及其对vsmcs增殖凋亡的影响》由会员上传分享，免费在线阅读，更多相关内容在工程资料-天天文库。
1、Jagged1真核表达质粒构建及其对VSMCs增殖凋亡的影响钱德慧，武晓静，姜洪，匡春燕，王逵，宋明宝，黄岚（400037重庆，第三军医大学附属新桥医院心内科）[摘要]目的 构建大鼠Jagged1真核表达质粒，分析Jagged1过表达对血管平滑肌细胞（VSMCs）增殖和凋亡的影响。方法 从带Jagged1全长基因质粒pBOS-SN3T中酶切获得目的基因，用载体pCMV-IRES-GFP构建重组质粒，转染至VSMCs。Westernblotting检测Jagged1蛋白表达变化，[3H]-TdR掺入法检测细胞增殖，流式细胞仪(FACS)检测细胞凋亡。结果 成功构建大鼠Jagged1真核表达载体并
2、转染至VSMCs，经G418筛选后转染率可达90%以上，转染上调了VSMCs的Jagged1蛋白水平约6倍。转染p-rJagged1后VSMCs的增殖能力明显降低（10265±1004vs.17584±1587cpm/well,n=5,P＜0.01），而凋亡明显增高（总凋亡率：32.35±2.86%vs.15.24±1.27%；早期凋亡率：22.47±2.54%vs.10.22±0.98%；n=5,P＜0.01）。结论 成功构建大鼠Jagged1真核表达质粒,过表达Jagged1可抑制VSMCs增殖并促进其凋亡。[关键词]平滑肌细胞;细胞增殖;细胞凋亡;Notch通路 [中图分类号]R363C
3、onstructionofaneukaryoticvectorexpressingJagged1geneanditseffectonproliferationandapoptosisofvascularsmoothmusclecellsQianDehui,WuXiaojing,JiangHong,KuangChunyan,WangKui,SongMingbao,HuangLan(InstituteofCardiovascularDiseasesofPLA,XinqiaoHospital,ThirdMilitaryMedicalUniversity,Chongqing400037,China)[
4、Abstract]ObjectiveToconstructarecombinanteukaryoticvectorexpressingratJagged1,andtoexploretheeffectofJagged1over-expressionontheproliferationandapoptosisofvascularsmoothmusclecells(VSMCs).MethodsThefragmentofJagged1genewasobtainedfrompBOS-SN3T(encodingratJagged1)bydigestion.Aneukaryoticvectorexpress
5、ingratJagged1(p-rJagged1)wasconstructedwithpCMV-IRES-GFP,andtransfectedintoVSMCs.TheexpressionofJagged1proteinlevelwasdetectedbyWesternblotting.TheproliferationofVSMCswasobservedbyassessingtheincorporationof3Hthymidine.TheapoptosisofVSMCswasanalyzedbyflowcytometry.Results Recombinantplasmidp-rJagged
6、1wasconstructedandtransfectedintoVSMCssuccessfully.Theefficiencyoftransfectionwasatleast90%.TheproteinlevelsofJagged1wereincreasedbyalmost6foldsaftertransfect..TheproliferationofVSMCswassignificantlylower(10265±1004vs.17584±1587cpm/well,n=5,P＜0.01),andtheapoptoticrateofVSMCswassignificantlyhigher(to
7、talrateofapoptosis:32.35±2.86%vs.15.24±1.27%;earlyrateofapoptosis:22.47±2.54%vs.10.22±0.98%;n=5,P＜0.01)inthep-rJagged1transfectedgroupcomparedwiththatintheuntransfectedgroup.ConclusionJagged1over-expr
