鈍頂螺旋藻表達載體的構(gòu)建及轉(zhuǎn)化研究.pdf

1、安徽農(nóng)業(yè)大學(xué)碩士學(xué)位論文鈍頂螺旋藻表達載體的構(gòu)建及轉(zhuǎn)化研究姓名：王芬申請學(xué)位級別：碩士專業(yè)：微生物學(xué)指導(dǎo)教師：唐欣昀20080601AbstractInthispaper，thetoleranceandsensitivitiesofSpirulinatodifferentantibioticsandLcanavaninesulphate(CS)wasexpoundedIntheexperiment，weextractedDNAantiC

2、SmutantA9candtransformeditintoA9strain，thenweacceptedsomeparameterstotransformSpirulinaAlso，weconstructedtheforeignvectorp215t—sppwhichcontainsSpirulinapromoterA9，andtransformeditintoSpirulinapromoterA9Thus，GreenFluoresc

3、entProtein(GFP)expressedsuccessfullyandstablyinSpirulinaThisresearchestablishedasystemforgenetictransformationoperationofSpirulina，andprovidedimportantreferencetotrans—genetechnologyofSpirulinaSpirulinawasextremelysensit

4、ivetoantibioticssuchasChlandStrThelethalconcentrationofChlandStrwere1Olxg‘ml～and31xg。ml。1inliquidcultivatingand121xg‘ml～and4pg‘ml?！甶nsolidcultivatingrespectivelyTheSpirulinawasrelativelysensitivetoAmpTheminimuminhibitory

5、concentration(MIC)ofAmpwaslOOpgml?！痠nliquidsituationand200jxg’ml～insolidsituationBut600lrtg‘ml‘1KmshowednoinhibitiononthegrowthofA9strainChl，StrandAmpmightbesuitableselectiongeneticmarkersfortheSpirulina，andtheselectionc

6、oncentrationswere12Itg。ml‘‘、41xg‘ml。1、2001xgml。1respectively，ExtractedDNAofA9cstrainwhichistheanti—CSmutantofA9strain，screenedby301xg‘ml—CS，thentransformeditintoA9strain，weacceptedatransformantandsomeparameterstotransfor

7、mSpirulina：culturedA9strainat24。Candtreatedwith2mmolL。1EDTAfor24hbeforetransformation；usedultrasonictreatmentwithpowerof300wfor70s，usednaturalmethodstoscreentransformantsInthisexperiment，bothoforiginalplasmidp2’15tandnew

8、constructedplasmidp215tsppwhichcontains舴genewereusedasvectors，useddifferentmethodoficebathandPEGfusionmethodseparatelytotransformSpirulinaA9strains，the肋genewereexpressedinSpirulinasuccessfullyWecomparedthetransformationr

9、ateofdifferentplasmidunderthesalTleconditionsandmethodsThedatashowsthat，comparedwithp215t，p215t—sppcallsignificantlyimprovetransformationrate(a=oo1)ThisprovedthatuseSpirulinapromotertoconstructvectorsCanimprovethetransfo

10、rmationrateWealsocomparedthetransformationrateofdifferentmethodsThedatashowsthatuse1％PEGconcentrationin30minCanobtainahighertransformationrate(105‰)Observedbyfluorescencemicroscopewith390nmasstimulationlight，partsofthetr