人脑源性神经营养因子和神经营养素3真核双表达载体的构建与鉴定-论文.pdf

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1、中国组织工程研究第侣卷第15朔2014—04—09出版ChineseJournalofTissueEngineeringResearchApril9，2014VoL18,No．15WWW．CRTER．org—I●●■■■■_·__一‘■■●●onStrUCtIonanaIdentifcationofbICIStrOnICe．uK。aryoti。ceXPress!■onVeCt■orOt，n■Umanb●raI■n-d■erI■Ved■neurotrophicfactorandneurOtrOphine·3LiBing—nan，LiWei-

2、dong，LinJun-tang，FengHui-genDepartmentofLifeSciencesandTechnology,XinxiangMedicalUniversity,Xinxiang453003,HenanProvince，ChinaAbstractBACKGROUND：Humanbrain—derivedneurotrophicfactor(BDNF)andneurotrophine．3(NT．3)areessentiaILiBing·nan．PhD，Lecturer,genesforcelIdiferentiatio

3、n．ViraIvectorhasbeenusednumerouslyinciinicaIpractice．butthesecurityistheDepartmentofLifeSciencesmostjmportantproblem．Eukaryoticexpressingvectorisawaylosolvethisquestion．andTechnology,Xinxiang0BJECTIVE：T0constructandidentifyplRES2-BDNF-NT-3bicistroniceukaryoticexpressionve

4、ctor．MedicalUniversity,Xinxiang453003。HenanProvince．METHODS：BDNFandNT．3geneswereobtainedfr0mthegenomicDNAofhumanperipheraIbloodChinamononuclearcelsbyPCR．TheBDNFcDNAfragmentwasinsertedintothemultiplecloningsitesofLiWei-dong，Studyingformaster’sdegree．DepartmentandOfLIfeScie

5、ncesandTechnology,XinxiangRESULTSANDC0NCLUSION：TheDNAsequencinganalysisdemonstratedthattheBDNFandNT．3wereMedicalUniversity,XinxiangexactlyconsistentwiththesequencerecordedintheGenBank．Enzymedigestionanalysisindicatedthat．inthe453003，HenanProvince。constructedbicistroniceuk

6、aryoticexpressionvectorPIRES2-BDNF．NT．3．BDNFbandwasobtainedbyEcoRlChinaIBamHIdigestion．IRES．NT-3fragmentwasobtainedbyBarnHI／NotIdigestion．andBDNF．1RES．NT．3wasLiBing-nanandLiWei-dongobtainedbyEcoRl／Notldigestion．RT．PCRandwesternblotanalysisshowedthat．aftertheHEK293cellscon

7、tributedequalytothisweretransfectedwithPIRES2-BDNF．NT．3．doublegenewasexpressedatthemRNAandproteinIeveI．workExperimentalfindingssuggestthat．bicistroniceukaryoticexpressionvectorofBDNFandNT一3genescanbesuccessfullyconstructedusingIRESsequence．Correspondingauthor：LiBing-nan．D

8、epartmentofLifeSubjectheadings：brain-derivedneurotrophicfactor；nervegrowthfactors；carrierprotein