1、北京化工大學(xué)碩士學(xué)位論文霉菌脂肪酶基因的克隆與表達姓名：江慧芳申請學(xué)位級別：碩士專業(yè)：生物化工指導(dǎo)教師：王雅琴20070613CLONINGANDEXPRESSIONOFLⅡ!ASEGENEFROMRH眨：DP嬲D霞l，z4ELipaseisatypicalenzymewhichcatalyzesthehydrolysisoflongchaintriglyceridesneyconstitutethemostimportantgroup

2、ofbiocatalystsforbiotechnologicalapplicationsLipasefromRhizopusoryzaehas13positionalspecificityitcanhydrolyzetriacylglyc2rolproducingfattyacidandmonoglyeeride(MG)，MGisanimportantnonionicsurfactants，widelyusedasemulsifier

3、inthefoodstuffandotherindustriesAsaresulttodaypeoplefocusontheconstructionofgenerecombinantstrainwithhigbenzymeactivitybygenemeasusesBesides，Thereisstillnouniformmethodtomeasuretheactivityoflipaseexactly，whileitisimporta

4、ntinreasearchingofcharactersandamplicationoflipasesInThisthesis，thegenewhichencodinglipaseROLfromRhizopusoryzaeWasamplifiedbyPCRandclonedintove貯torpUCmTWassavedinourlabTheRecombinantplasmidIsnamedpGMTROLandIdesignedprime

5、rpair4and42toclonethelipasegeneROLfrompGMTROLseparatedconnectwithpET28aandpETllcTherecombinantplasmidarenamedpET28aROLandpET11cROLandpET28aROLWastransferredintoEcoliBL21andpET11cROLWastransferredintoEcoliOrigamiDE3Expres

6、sionRecombinantstrainswereobtainedseparatedbyKanaandAmpplatesscreening，theExpressionRecombinantstrainswerefartherfilteredbystrainPCR，andTheengineeredstrainwasexpressedthegeneROLbytheinductionofIPTG’11leexpressionproducts

7、ofROLgenewasanalysisbySDSPAGEtheresultindicatedthatROLgeneWasfunctionallyexpressedinEcoliBL21andOrigamiDE3Recombinantlipasehadamolecularmassof35kDOntheothersideThisthesisdiscussesthreeassaysincommonuse，includingthespectr