高溫α-淀粉酶產(chǎn)生菌的遺傳改良及發(fā)酵工藝優(yōu)化.pdf

1、江南大學碩士學位論文高溫α淀粉酶產(chǎn)生菌的遺傳改良及發(fā)酵工藝優(yōu)化姓名：楊艷華申請學位級別：碩士專業(yè)：發(fā)酵工程指導教師：王正祥20090301AbstractAbstractThethermostableⅨ一amylaseisoneofthemostimportantcommercialenzymeswithanumberofapplications，includingitsuseinstarchprocessing，sugar，texti

2、leandpaperindustriesUptothepresent，Bacilluslicheniformisistheonlyindustrialstrainforproductionofthermostable伐amylaseThisstudyhasinvestigatedthegeneticimprovementandfermentationprocessoptimizingoftheindustrialstrainBacill

3、uslicheniformis，hopingforincreasingtheexpressionlevelofthethermostableaamylaseBasedontheoptimizationtheproteinexpressionsystemofthethermostable僅amylaseproducingstrain丑licheniformistheaimofthepresentworkistoperformthegene

4、ticimprovementof丑licheniformisbygeneticengineeringInthiswork，a600bpfullgenefragmentofdegQthatamplifiedfromthegenomicDNAofB1icheniformisbyPCRmethod，WasdigestedwitllBamHIandinsertedintotheBglIIsiteofpHYP43producedarecombin

5、antplasmidpHYP43degQTherecombinantplasmidpHYP43一degQWastransformedinto最licheniformisATCC14580andB0204byelectroporationmethod，resultedinrecombinantstrainsATCCl4580(pHYP43一degQ)and(pHYP43一degQ)，respectivelyTheresultsofshak

6、ingflaskfermentationindicatedthatthe僅一amylaseproductionlevelbystrainATCCl4580(pHY—P43degQ)Was354U／ml，whichisalmost30foldsoftheoriginalstrain(12U／m1)，andthatofstrainB0204(pHYP43degQ)increased1518％Afurtherstudyhasdonetoinv

7、estigateandoptimizethefermentationprocessoftherecombinantstrainB0204(pHYP43一degQ)ThestudiesonthephysiologicalcharactersofstrainB0204(pHY—P43一degQ)showedthatitWasabletoutilizeglucose，lactose，sucrose，glycerol，galactitol，ga

8、lactose，sodiumgluconateandgluconate，andCangrowatatemperaturerangeof34500CandatapHrangeof5578晰thitsoptimalgrowthconditionsoftemperatureof42oCandpHof65，respectivelyThisstudyhasinitiallyconfirmedthepropermediumandfermentati

9、onprocess，andtheoptimizedshakingflaskfermentationconditionswere：beanpower25％，lactose4％，P04。40mmol／L，pH60，inoculation10％Aftertheoptimizationofthefermentation，thethermostable0【amylaseproducedbytherecombinantstrainB0204(pHY

10、P43degQ)wasincreased715％Ontheoptimizedfermentationcondition，therecombinantstrainB0204(pHYP43一degQ)produced28％thermostableaamylasemorethanthatofstrainB0204KeyⅥords：Thermostable僅一amylase，Expression，BacilluslicheniformisFer