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20104452121126JOURNALOFGANSUAGRICULTURALUNIVERSITYB2,,,,(,,2,730070):B2,B2.:B210%,pH7.5,1000Lh-1,200rmin-1;20h;36h.,.:B2;;;:TQ929:A:100324315(2010)0220121206:(19842),,,.:,,,.E2mail:chenxiurong@gsau.edu.cn:(GNSW22005210);555.:2009204228;:2009205229FermentationtechniquesonBacillussubtilisB2andqualitydeterminationofproductsJINGZhuo2qiong,LIMei2li,CHENXiu2rong,YANGCheng2de,XUELi(CollegeofPrataculture,GansuAgriculturalUniversity,KeyLaboratoryofGrasslandEcosystem,MinistryofEducation,Sino2U.SCentersforGrazinglandEcosystemSustainability,Lanzhou730070,China)Abstract:InordertoconductthetrainingofBacillussubtilisB2fromtheseedfermentationtofermen2tationbroth,orthogonalexperimentswereadoptedtostudytheoptimizationoffermentationB2,fermenta2tiontimeofseedtanksandbrothandthequalitydeterminationmethods.Theresultsshowedthatfermenta2tionconditionsfortheoptimalcombinationwasinoculum10%,pH7.5,ventilation1000Lh-1,rotatespeed200rmin-1;theseedtankfermentedfor20h;theoptimumharvesttimewas36hforthebroth.Thequalityoflivecapacityforthefermentablewasthemaintargets,assistedturbidimetrydetection.Keywords:BacillussubtilisB2;fermentationtechnique;orthogonalexperiment;detection[1].,[2].(Bacillussubtilis),,,,[3],,,,,[4].,,(Stagonosporacurtisii)[5](Rhizocto2niacerealis)[6](Cladosporiumful2vum)[7](Fusariurngraminerarum)(Rhizoctoniaspp.)[8](Alternariaal2ternata)(Rhizoctoniasolani)(Fusariumoxysporumf.sp.vasinfectum)[9],,2010,,,,,,[10].B2,,[11].,,[12].,,B2,.11.11.1.1B2.1.1.2(NA):3g,5g,16g,1000mL,B2;(NB):3g,5g,2.5g,1000mL,B2;(1%,0.8%,0.5%)[11];PDA:.1.1.3S2000UV/VisAL104HH1.5PlusBLOF26005AGBN5LBLOF26010ASBN10LKR25i.1.21.2.1(121,30min)()()pH(121,20min)(NB24hB2).1.2.2,200Lh-1,200rmin-1,pH8,5%,60%,28,4h1,;[13],.1.2.31.2.2[11],2810LpH(1).,10[14],y()=10x,,x.1L9(34)Tab.1ThefactortableoftheorthogonaldesignL9(34)/%pH/(Lh-1)/(rmin-1)157.52002002108.04003003158.56004001.2.4pH1.2.3,pH2.1.2.3.1.2.5[13],1.2.4.1.31.3.110mL15min(12000rmin-1),(V),(10-V)/10[15].1.3.2,,600mm(D600nm).1.3.31mL,10-710-810-9(),0.1mLNA.282d,.(cfumL-1)=/(mL)D600nm,,.2212:B21.4ExcelDPS.22.11,B2,,,04h,,B2;424h,20h2.86108cfumL-1,,;2444h,108,;48h,,.,B2,2024hB2,,,,20h.1B2Fig.1ThegrowthcurveofB22.2,2,4,36h,,36h40h44h,3(P0.05),36h,,,,36h.2Fig.2Theeffectofinhibitoryonplantpathogenicfungusinfermentationliquid2.32,,95,5.4601010cfumL-1,2,1.0071010cfumL-1,1.1131010cfumL-14.6631010cfumL-1,54.91.17,,510%,2L9(34)Tab.2TheschemeandresultoftheorthogonaltestL9(34)/%pH/(Lh-1)/(rmin-1)F0.05F0.01157.520020010.556cB258.040030010.009gG358.560040010.429dC4107.540040010.119fEF5108.060020010.736aA6108.520030010.274eD7157.560030010.664bA8158.020040010.046gFG9158.540020010.140fE3212010pH8,600Lh-1,200rmin-1.,,.,.3,pH4,,0.520,,,.,0.093,.3,3(1)43Tab.3ThecompletelyrandommodelresultanalysisoftheorthogonaltestRR’10.28310.3760.0930.084pH10.26410.4460.1830.16410.09010.6090.5200.46810.19810.4770.2790.252,(4),pH7.5,3pH8,pH.4SSRTab.4TheSSRtestofsignificantdifferencesbetweenthelevelofthefourfactorsF0.05F0.01pHF0.05F0.01F0.05F0.01F0.05F0.01110.331bA10.446aA10.292bB10.477aA210.377aA10.264bB10.089cC10.316bB310.283cB10.281bB10.609aA10.198cC2.4pH5,21(P0.05),1.631011cfumL-1,pH7.5pH8B2,,pH7.5.5pHTab.5ThevarianceanalysisofoptimizationofthepH/%pH/(Lh-1)/(rmin-1)F0.05F0.01110860020010.350bA2107.560020011.200aA(6),((P0.01)),3,1.731012cfumL-1;121.631011cfumL-13.421011cfumL-1,CK(2.241010cfumL-1)(P0.01),10%,pH7.5,1000Lh-1,200rmin-1.6Tab.6Thevarianceanalysisofoptimizationoftheventilation/%pH/(Lh-1)/(rmin-1)F0.05F0.011107.560020011.202cC2107.580020011.533bB3107.5100020012.237aACK108.060020010.350dD2.512200Lh-1,200rmin-1,pH8,5%,,,(1000Lh-1,200rmin-1,pH7.5,10%),(3),,4212:B23Fig.3Theeffectofinhibitoryonplantpathogenicfungusinfermentationliquid4,3644h.36h3.8172.6502.2173.750,21.4070.8671.1981.1502.7123.0571.1513.261,,.,36hB2.2.67,39,571369482,D600nm571639842,,,578349126,2,.7Tab.7ThemethodofqualitydetectioninfermentationliquidF0.05F0.01D600nmF0.05F0.01F0.05F0.0155.460aA39.985aA0.079aA74.663bA38.727aA0.068bAB13.590cB30.545bB0.051cdC32.667dC27.596bcB0.059bcdBC61.877eD29.201bcB0.048dC91.357fE27.479cBC0.052cdC41.317fEF23.919dC0.058bcdBC81.113gFG24.141dC0.061bcBC21.007gG18.373eD0.048dC(8),4D600nm4321,.,D600nm,,,,,,(3d);,,,,,,,,,.8Tab.8Thevarianceanalysisofthemethodofqualitydetectioninfermentationliquid/%pH/(Lh-1)/(rmin-1)F0.05F0.01D600nmF0.05F0.01/cmF0.05F0.0111086002000.224dA11.193dD1.298dC2107.56002001.625cB14.221cC1.854cB3107.58002003.415bBC17.601bB2.375bB4107.5100020017.283aC19.830aA3.817aA521201033.1.,,,,[16].,,,(20h),,.3.2,[17],B2.,2,,,,,,,,,B2.3.3pH,B210%pH7.51000Lh-1200rmin-120h,36h,(1.731012cfumL-1),B2pH8(2.971010cfumL-1),,pH,pH,,,,pH,pH,,;,,pH7.5,pH,[11,18]..,,,,,,,[19221],.,,,,,.,,.,,,,.[1],,,.[M].:,2000[2]CookRJ,BruckartWL,CoulsonJR,etal.Safetyofmicroorgnismsintendedforpestandplantdiseasecon2torl:aframeworkforscientificevaluation[J].Biologi2calControl,1996,7:3332351[3]MonicaLE,ElizabethADJ,WilliamEBJ.etal.Via2bilityandstabilityofbiologicalcontrolagentsoncot2tonandsnapbeanseeds[J].PestManagementScience,2001,57(8):6