百脈根中脫氧葡萄糖篩選表達(dá)載體的構(gòu)建和轉(zhuǎn)錄因子CBF1的轉(zhuǎn)化研究.pdf

1、東北師范大學(xué)碩士學(xué)位論文百脈根中脫氧葡萄糖篩選表達(dá)載體的構(gòu)建和轉(zhuǎn)錄因子CBF1的轉(zhuǎn)化研究姓名：郭喜英申請學(xué)位級別：碩士專業(yè)：植物學(xué)指導(dǎo)教師：王興智邢少辰20070501AbstractAbioticstress，especiallylowtemperature，droughtandhighsalt，hasbeenanegativefactorintheagriculturalactivitiesoverpastyearsandbecom

2、emoreseriousduetothechangeofglobalclimateItisveryimportanttoimprovetheplanttoleranceagainstvariousabioticstressesbymeansofplantmoleculartechnologyinagricultureWiththediscoveryofcoldregulatedgenes，manyresultshavebeenrepor

3、tedthatthosegenesweretransferredintotheplantssuccessfullyandconsequentlyimprovedtheabilityoftoleranceagainstlowtemperatureMostofthesegenesareregulatedbyCBFtranscriptionfactorssOitisanovelchanneltointroducethetranscriptio

4、nfactorgenesintoplantstoimprovecropcoldresistanceInthisstudythedeoxyglucoseselectionsystemwasfirstlysetupincrowtoe(Lotuscorniculatus)，andthentheplantexpressionvectorscarryingtranscriptionfactorCBFlweTeconstructedbasedonb

5、otllherbicideanddeoxyglucoseselectionsystemsrespectivelyTransformationofCBFlgenewasperformedbytheAgrobacteriummediatedCO—cultivationmethodThemainresultsareasfollows1Obtainmentofthegenes：AccordingtotheDNAsequenceoftranscr

6、iptionfactorCBFlgenepublishedintheGenBankapairofprimerWasdesignedandsynthesizedUsingthetotalgenomicDNAofArabidopsisthalianaseedlingastemplates，thefulllengthofCBFlgeneWasamplifiedbyPCRandclonedintoTEasyvectorDNAsequencing

7、resultindicatedthatthetargetfragmentshowed100％identitytothegenesequenceintheGenBankTheselectiongeneDOGRlWasobtainedbydigestionofpBinAR：：DOG81vectorwithrestrictionenzymesSa／IandEcoRI2Constructionofplantexpressionvectors：T

8、hreeofplantexpressionvectors，includingpCAMBIA3300一CBFl，pCAMBIA5300andpCAMBIA5300CBFl，areconstructedAlltheCBFlandDOG“1genesinthosevectorsaredrovebythesamepromoterCaMV35S3Establishmentofthedeoxyglucoseselectionsystemincrow

9、toe：Thesystemhasbeenbuiltthroughthescreeningofdifferent2deoxyglucoseconcentrationWefoundthatasmallpartofcrowtoecotyledonsdifferentiatedatfwstonthemediumwith600mg，L2deoxyglucose，butdiedgraduallyafterwardsWhentheconcentrat

10、ionof2deoxyglucoseishigherthan1000mg／L，nocrowtoecotyledonscouldbesurvivedtodifferentiate4Obtainmentofthetransgeniccrowtoeanditsinvestigation：Theratesoftransformationbasedonbothdeoxyglucoseselectionandherbicideselectionar

11、esimilarUnderthenormalgrowthconditiontheprolinecontentsintransgenicplantsderivedfrombothselectionsystemsarelitterhigherthanthatofuntransformedplants，andsynchronousincreaseddramaticallyafter4“Ctreatmentreachedthemaximumva