CpG重組質粒的體外篩選及其在豬體內的安全性研究.pdf

1、中國農業(yè)科學院碩士學位論文CpG重組質粒的體外篩選及其在豬體內的安全性研究姓名：嚴彩紅申請學位級別：碩士專業(yè)：預防獸醫(yī)學指導教師：朱鴻飛20050601AbStractTheSPFswineperipheralbloodmononuclcarcells(PBMCs)byFicollgradientcentrifugationprocedure，werepreparedandstimulatedbyrecombinantpcDNAKCpG

2、一1，pcDNAKCpG一2peDNAKCpG一3andpcDNAKCpG一4withfourdoses(125pg，25／tg，50lag，10O煺)respectivelyTheeffectofthelymphocytetransformationwasdetectedby～rTTassayItisshowedthatthefourplasmidshaddifferentextentimmtmostimulatoryeffect；t

3、hestimulateindexofthe10pgofpeDNAKCpG4is63Afterelaborateanalysis，theconclusionwasdrawnthatthepeDNAKCpG2hadthemostimmtmostimulatoryactivityonswinebecauseithadagoodimmunostimulatoryeffectevenatlowdosesTheswineinjectedwith50

4、0pgofpcDNAKCpG2withthemostintmunstimulatmyactivitywereslaughteredtoextractthegDNAofheartliverspleen，lung，kidneybloodandmuscleoftheinjectionsiteat4h，4d，8d15dand30dafterinjectionThenthedistributionandpersistenceoftheplasmi

5、dinthetissuesweredetectedbyPCRusingspecificprimersItisshowedthatpeDNAKCpG2wasdetectedinmuscleat4hafterinjection；FourdayslaterthefragmentofplasmidwasamplifiedfromthebloodandmuscleofonepigbyPCRTheplasmidcouldexistinorgansn

6、omorethan8daysafterinjectionOnthebaseofaboveresearch，thegDNAofPCRpositiveandNO9pigat30dafter螄ectionwasidentifiedbysouthernblottingwithaDIGlabeledprobetodeterminewhethertheplasmidhadintegratedintohostchromosomeornotResult

7、sshowedthattheplasmiddidnotintegrateintothegDNAofswineGenerally，peDNAKCpG2hasthemostimmunostimulatroyeffectonswine；thedistributionandpersistenceoftheplasmidcouldexistinorgansnomorethan8daysafterinjeetion；anditwouldn’tint