FXR1P相互作用RNA的文庫篩選.pdf

1、南華大學碩士學位論文FXR1P相互作用RNA的文庫篩選姓名：喻長順申請學位級別：碩士專業(yè)：生物化學與分子生物學指導教師：何淑雅20070501IVAbstractObjective:FragileXsyndrome(FXS)isoneofthemostcommonfmofinheritedcognitiveimpairmentwhichhaveadiversityofclinicalmanifestation.Themutationof

2、FMR1causesFXS.FXR1agenethatsharesgreatsimilaritywithFMR1(FragileXrelatedgene1)isahousekeepinggenethatplaysanimptantroleinthedevelopmentprocessofnervoussystemmuscularsystem.FXR1P(FragileXrelatedprotein1)thetranslationalpr

3、oductofFXR1isakindofRNAbindingproteinbutourknowledgeaboutFXR1PtargetingRNAsisextremelylimited.TherefewescreenedfromahumanbraingyrushippocampipRH3cDNAlibraryfFXR1PinteractingRNAsindertoenrichourknowledgeaboutthefunctionof

4、FXR1.Methods:ConstructhumanbraingyrushippocampipRH3cDNAlibrary.TheFXR1PwholelongcodingvectpYESTrp3FXR1wasintroducedintohoststrainL40ura3pHybLexZeoMS2；ThenthetoxicityselfactivationofpYESTrp3FXR1weretested；Thehumanbrainlib

5、rarywasscreeningbyyeasttransfmation；TheplasswereextractedfromtheobtainedpositiveyeastclonesreintroducedintoyeaststrainL40ura3pHybLexZeoMS2pYESTrp3FXR1onebyonesoastovalidatetheRNAproteininteractions；Thefragmentsofidentifi

6、edpositiveclonesweresequencedanalyzedwithBLASTinNCBI.Results:ThehumanbraingyrushippocampipRH3cDNAlibrarywassuccessfullyconstructedTheintroductionofpYESTrp3FXR1washarmlesstothehoststraintherewasnoselfactivation56strainsof

7、positiveresultswereobtainedthroughyeastthreehybridscreeningnineofthemwereverifiedbyonetooneyeastthreehybridsystemanalysisThenineclonesweresequencedthenconfmedbyBLASTanalysisfourofthemprovedtobeedcrectlynamelyHomosapienst

8、ubulinalphaubiquitous(KALPHA1)、HomosapiensIQmotifcontainingE（IQCE）、Homosapiensdipeptidylpeptidase7（DPP7）、Homosapiensleukocytereceptcluster（LRC）member4whilefiveofthemwereedreversely.OurfindingsmaycastanewlightonFXR1Pfunct