利用單酶切cDNA-AFLP技術分離玉米對生性狀相關基因差異表達片段.pdf

1、安徽農(nóng)業(yè)大學碩士學位論文利用單酶切cDNAAFLP技術分離玉米對生性狀相關基因差異表達片段姓名：李蕾娜申請學位級別：碩士專業(yè)：生物物理學指導教師：程備久朱蘇文20050601AbstractMaizewithoppositecharacterisbothagoodgermplasmresomceforhighyieldbreeding，andapreciousvariationforplantdevelopmentresearchIno

2、rdertostudythemolecularandregulationmechanismdeeply，inthisstudy，basedonconstructingsuitableNILs(nearisogeneticline)oftotalRNApoolsofoppositemaizeandmutualmaize，singleendonucleasedigestioncDNA—AFLPtechnologywasappliedtosc

3、reen，isolationandbioinformaticsanalyzethegenedifferentiallyexpressedfragmentsrelatedtotheoppositecharacterofmaizeTheresultswereasfollow1nlequalityandintegralityoftotalRNAextractedWithTrizolmethodweregoodandweresuitableto

4、constructthetotalRNApoolsofoppositemaizeandmutualmaizeequivalently2ThelengthofDouble—strandcDNAretransferedbySMARTTMPCRcDNASynthesisKitfromeachpoolwas100～1000bpItsintegralitywasfine3Thelengthofdigestedfragmentbyaendonucl

5、ease—Sau3AI，whichcanrecognizea4bpsequece，wasl00～1000bpItcouldaccordwiththeAFLPtechnology4，Theligationproductwaspre—amplifiedtogetnumbersofcopies，aftertheendonucleasedigestionfragmentwaslinkedtotheartificialadapter5Dilute

6、dpre—amplifiedproductwassystematicallyselective—amplifiedAimprovedPCRmethodTOUCHDOWNPCRprogramwasadoptedtogetpeculiarandhighdensitycopies6Combined、Ⅳithnreadenatured6％PAGEandsilverstain29differentiallyexpressedfragmentsha

7、dbeenobtained：23fragmentsenhancedexpressed，6fragmentsrestrainedexpressedinoppositemaize75enhancedexpressedfragmentsofoppositemaizewereclonedandsequenced，includingS”m、Snqc、STT《、S”《、So_‰Thehomologyanalysisshowedthatthese5f

8、ragmentshadveryhighhomologies、ⅣitllthemaizemRNAthathadebeenseparatedS丌GChad80％homologywithcytochromeb5geneofNtabacumScf1【，hadover90％homologywith5tublingeneofZmays，Eleusineindica，Hordeumvulgare，OryzasativaandSetariaviridi

9、sThisstudyfirstappliedthesingleendonucleasedigestioncDNA—AFLPtoanalyzethedifferentiallyexpressedgenesinmaize，andmadeaseriesofimprovementtothetechnologytofmdsomedifferentiallyexpressedfragmentsrelatedtooppositecharacterin

10、maizeFurtherresearchcarlbedonewiththeobtainedsequencestomakeprobesormolecularmarkerstolocate，clonetheoppositegene，andevenstudythegeneexpressioncharactersKeywords：Maizeoppositecharacterdifferentialexpressionsingleendonucl