果蠅microRNA-184功能的初步研究.pdf

1、東南大學(xué)碩士學(xué)位論文果蠅micrNA184功能的初步研究姓名：胡江波申請學(xué)位級別：碩士專業(yè)：生物化學(xué)與分子生物學(xué)指導(dǎo)教師：袁榴娣20080530英文摘要TheprimaryresearchofmicroRNA184functionsinDrosophilamelanogasterGraduateStudent：HuJiangboSupervisor：YuanLiudiKeyLaboratoryofDevelopmentalGenesan

2、dHumanDisease，?！疢inistryofEducationSoutheastUniversityABSTRACTInrecentyears，evidencehasaccumulatedthatsmallnoncodingRNAsarealsousedinaconservedmannertoregulatekeydevelopmentaleventsAtleastfourclassesofregulatorysmallnon—

3、codingRNAshavebeendescribed，includingmicroRNAs(miRNAs)，shortinterferingRNAs(siRNA)，repeatassociatedsmallinterferingRNAs(rasiRHAs)andpiwi—interactingRNAs(piRNAs)[62631AmongthesesmallRNAs，miRNAsarethemostphylogeneticallyc‘

4、onservedandfunctionposttraascriptionallytegulatemanyphysiologicprocesses，includingembryonicdevelopmentmig184isamiRNAconservedinDmelanogasterandDpseudoobscuramouse，humanItexpressesatallstagesofDmelanogasterTheexpressionle

5、velincreasesprogressivelyasindividualdevelopementmiR一184displaysexpressioninthemesoderm，anteriorendoderm，andposteriorendodermofgermbandextendingembryosbyinsituhybridizationtonascentmiRNAtranscripts[141Inordertostudytheim

6、portantregulationroleofmiR184duringtheDmelanogastergrowth，WeconstructedthetransgeneDrosophilaofmiR184WhereafleroverexpressionofmiR184wasfinishedbyusingofdifferentGal4toolflies，atthesametimewefinishedthenonisotopicdetecti

7、onofoverexpressionofmiRI84usingofNorthernblotwithdigoxigeninlabledRNAprobesLatemesodermexpressionusingthe24BGal4drivershowedinterestingresultsDrosophilacoulddevelopmenttothirdinstarlaval，butstayedatthisstageandcouldnotfu

8、rtherdevelopmenttopupaAtthesametime，thesizeofthirdinstarlavalbecamesmallerobvious；themouthhooksbacamemalformedWepresumeithasarelationwithecdysonesignalpathwayThenwe燈iedecdysonefeedingexperimentsEcdysonefailedtescusetheph

9、enotypeWealsostudymiR184loss—offunctiontobuildmiR184nullmutantWeobtainedaPelementstrainfromBloomingtonWeattemptedtogetmiR184nullmutantusingoflocalmutagenesisimprecisePelementexcisionInordertolookforthepossiblelytargetgen

10、eofmiR184，weattemptedtofilterthepossiblelytargetgenesofmiR184onthecelllevelWecotansfectionrecombineplasmidof3’UTRandpAc—miR184intoDrosophilaS2cellAccordingtotheresultsofrelativeluciferaseactivitythatwasmeasured，weanalysi

11、stherelationbetweenmiR184andthe3UTRoftargetgenesTheresultsshowedthattheluciteraSeexpressionlevelofwhichconnectedthe3’UTRofCG812l、CG6583、CGl0217reducedsignificantlyThisprovidescredibledataforsearchingoutthetruetargetgenes