重復(fù)多色熒光原位雜交技術(shù)的建立及其在食管癌中的應(yīng)用.pdf

1、石河子大學(xué)碩士學(xué)位論文重復(fù)多色熒光原位雜交技術(shù)的建立及其在食管癌中的應(yīng)用姓名：楊壹羚申請學(xué)位級別：碩士專業(yè)：病理學(xué)與病理生理學(xué)指導(dǎo)教師：李鋒王明榮20050501EstablishmentofRepetitiveMulticolourFluorescenceinSituHybridizationandStudyinEsophagealSquamousCeUCarcinomasMDStudent：YangYiLingAdvisor：Pro

2、fessorLiFengProfessorWangMingRongObjective：NumericalandstructuralchromosomeaberrationsarethedefiningcharacteristicofhumancancerInagenomewidesearchformolecularalterationsunderlyingtumorigenesisinesophagealsquamouscellcarc

3、inoma(ESCC)，thepatternsofaberrationsatchromosomallevelmightbeallessentialguide，ashasbeenshownformanyotherhumanneoplasms，Duetotechnicalproblems，thechromosomemorphologyofsolidtumorsingeneral，andepithelialtumorsinparticular

4、isoftenpoorandasaconsequence，theputativegeneticeventsthatmightbeimportantinESCCremainundefinedTounderstandthechromosomalaberrationsofESCC，twoinvestigationswerecarriedoutinthisproject：establishmentofrepetitivemulticolourf

5、luorescenceinsituhybridization(RM—FISH)andprimaryculturetechnique，andjdentificationofchromosomalimbala／icesinvolvedinESCC’eelllinesMethods：The24microdlssectedchromosomes(22paksofautosomesplusthetwoSexchromosomes)weresubj

6、ectedtoDOP—PCRandthePCRproductsweretaggedwithdifferentcombinationoffluoresceinisothiocyanate(FITC)，Cy3andCy5Fourpoolsof6colourpaintingprobesweredesigned，andinsituhybridizationwasperformedforfourtimesonthesamespecimenFISH

7、imageswerecapturedwithanepifiuorescencemicroscopeequippedwithaCCDcameraand4positionfilterwheelByPdvlFISHcombiningwithinvertedDAPIbanding，weanalyzedthechromosomalalterationof3ESCC’ceillinesThekaryotypedescriptionwasaccord

8、ingtotheInternationalSystemforHumanCytogeneficNomenclature(ISCNl995)Tumorspecimenswereobtainedatthetimeofsurgeryfrom15patientswithESCCtoprocessedmetaphasepreparationandanalyzedmetaphasepreparationofaprimaryESCCbyRM—FISH，

9、Results：1)WedevelopedamodifiedrepetitivemulticolourFISHprotoc012)RMFISHtechniquecombiningwithinverted46一diamidino2一phenylindole(DAPI)bandingidentifiedthekaryotypeinthreeESCC’celllinesandfoundanumberofchromosomeaberration

10、sincludingdeleting，inserting，translocationandmarkchromosomesChromosomes1，5，11havehigherprobabilitiesofundergoingstructuralandnumericalaberrations；weobservedlossof3p，gainof3qandlossofYinallcelllines；iOq)intwocelllines(KYS