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562000,Vol.20,No.2李民陈常庆(,200233)重组大肠杆菌的高密度发酵是提高基因工程产品产量的一个非常有效的手段,是现代发酵工程研究的一个热点本文就高密度发酵中影响重组大肠杆菌发酵产率的几个因素,包括宿主菌培养基培养条件补料方法以及高密度发酵过程中存在的问题和对策加以讨论,着重探讨了高密度下大肠杆菌产生的有害代谢副产物)))乙酸的产生机制抑制作用机理,以及控制乙酸积累的技术方法重组大肠杆菌高密度发酵补料分批培养比生产率乙酸DNA,,5-15%[1],20032500,10%[2](Highcell-densityculture,HCDC),,,(),,,,,pH,(TNFA),-3(IL-3),-2A(BMP-2A),1111CHONSPFeMgK,1g/L(L-1):0177gNH4Cl,01125gKH2PO4,1715mgMgSO4#7H2O,715mgK2SO4,0164mgFeSO4#7H2O,014mgCaCl2[3],,,,,,;,,;,,,,,,,,;,,,:(50g/L)(3g/L)Fe2+(1115g/L)Mg2+(817g/L)PO43-(10g/L)Zn2+(01038g/L)[4],,AllenLuli[5],(13h)79g(DCW)/L,Jung[6]C/N,INF-B,C/N,Tsai[7],IL-2,19%2%112,,26DOI10.13523/j.cb.20000206E.coliKPLPRtactrpIpplac,,,,,,,,,,BMP-2A[8]IL-2[9],3)4h(BMP,2178g/L;IL-2,1102g/L),,tacIPTG8)10h[10],113,,,,;,,,;,,,E.coliDH1YK537KS476proapoA-I2)3,20%)100%[10]Luli[11],E.coliJM105E.coliB(2g/L),(30g/L),MC106010g(DCW)/LWalle[28]E.coliBL21(DE3)JM109,1/4,25%,114[12],,[13],EngbergNord-storm[14],,,;Jones[15],8,(pBR322pMB9),4-5,2,Riesen-berg[4]400g(DCW)/L,Markl[16]200g(DCW)/L,25%3Lm,1Lm;,,E.coliW3110[16]-3-[17],:174g(DCW)/L17514g(DCW)/L11[1]/[gDCW/L]MC1061OD525=1201108g/L-1K1220600mg/LMC1061111175g/L-1AAM-768516g/L-2M52485915112g/LJE5505261@109U/LA-1TG160515@108U/LA-1TG195.52185@106U/L-2M52485915112g/L-2HW22-2OD680=90--2MM294OD680=75313g/LJM10340338mg/LW3110OD600=100219g/LHB101145610U/mgX909256mg/L27/[gDCW/L]B-JM103844600U/ODA-AM-768516g/LA-B-KA197771912g/LMini-RV308501104g/LA-B-TG195152185@106U/LB-C6006316100U/gXL1-Blue101148112g/LXL1-Blue175146515g/LW124163413g/LAT24713646g/LLFO3301102116@105U/g-2A[8]YK537OD600=532178g/L-3[21]YK537OD600=53313g/L[25]YK537OD600=1206g/L[36]YK537OD600=605112g/LC-[37]DH5A10111@1010IU/L3,,,/0,[8,11,18],,:[6,19][8,20,21][6,22,23],,,,,Pan[9],,120OD525;Jung[6],46g(DCW)/L,17mg/g[8,21],[8],YK537/pDH-B2m(BMP-2A),53OD600,BMP-2A2178g/L,,,,,,,,,,,22,,(),,,pHpH,,,pH,,,CER(CER),,DO-stat,,284,,,[1,5,8,11,18,24]411,,,,,HPLC,[24,25]RP-HPLC,YK537/pSB-TK,,,412,,Han[26],,,,ATPNADH2,(Pta)(Ack),AEL-Mansi[27]Luli[11]:pH(CH3COO-)(CH3COOH),,(pH715)CH3COO-H+,pH,pH-$pH,,,413(1),LuliStrohl[11],3E.coliBE.coliK[28],E.coliBJM105,MC1060,,,E.coliB(PtaAck)Neway[29]E.coliMM-294()IL-2,,,IL-2(2),[26]Kleman[30]pH,W3100015g/LpH610,6g/L,pH71512g/L[24]Han[26](),,(3)(L),,,,[26],012h-1Riesen-berg[31]0111h-1,,110gDCW/LL,L(4)37e26)30e,[32],E.coliKS467[32]ProapoA-I37e30e,10g/L5g/L(5)[33],Lee[33],29(6),,pH,DO,L,OUR,CER,,,,A,pH[6,30]pH,pH,pHpH,B,[27,30],,,,,,,,C,DO-stat[8,34],,,,DO-stat,DO-stat/0DO-statDO,,,5,,,[8]BMP-2A,32%Shimizu[35]IPTG,0102mM-2mM6,,()(pHCO2)(//)(),,,,;;;,,,[1]Lee,S.Y.,TIBTECH,1996,14:98-105.[2],,,,,1996[3]ReilingH.E.,LaurialH.,andFiechterA.,JBiotech,1985,2(3):191-206.[4]RiesenbergD.,Curr.Opin.Biotechnol.,1991,2:380-384.[5]AllenB.,andLuliGW.,Biopharm.Manuf.,1987,1:38-41.[6]JungG.,DenefleP.,BecquartJ.etal.,Ann.InstPasteurMi-crobio,1988,139:129-146.[7]TsaiLB.,MannM.,MorrisF.etal.,JIndMicrobio,1987,2:181-187.[8],,,,1998,14(3):270-275.[9]JinSD.,ChungBH.,HwangYB.etal.,JournalofFermenBioeng,1992,74(3):196-198.[10]OhtaK.,ShibuT.,MorimotoY.etal.,JournalofFermenBio-eng,1993,75(2):155-157.[11]LuliGW.,andStrohlWR.,ApplandEnvirMicrobio,1990,56(4):1004-1011.[12]MoserDR.,andCampbellJL.,JBacteriol,1983,154(2):809-818.[13]Uhlin,B.,andNordstromK.,Plasmid,1979,1:1-8.[14]EngbergB.,andNordstromK,JBacteriol,1975,123(1):17930-186.[15]JonesIM.,PrimroseSB.,RobinsonA.etal.,MolGenGenet,1980,180(3):579-584.[16]MarklH.,ZenneckC.,DubachA.etal.,ApplMicrobioBiotechnol,1993,39:48-52.[17]LeeSY.,andChangHN.,JEnvironPolymerDegrad,1994,2:169-174.[18]DoelleHE.,EwingsKN.,andHollywoodNW.,AdvBiochemEng,1982,23:1-35.[19]PanJG.,RheeJS.,andLebeaultJM.,BiotechnolLett,1987,9:89-94.[20]PaalmeT.,TilsmaK.,KahruA.etal.,BiotechnolBioeng,1990,35:312-319.[21],,,,1998,.[22]YeeL.,andChangBH.,BiotechnolLett,1993,15:971-974.[23]LeeJH.,ChoiYH.,KangSK.,BiotechnolLett,1989,11:695-698.[24],,,,1998,28(3):30-32.[25],,,,1999,29(1):25-28.[26]HanK.,LimHC.,andHongJ.,BiotechnolBioeng,1992,39:663-671.[27]EL-MansiEMT.,andHolmsWH.,JGenMicrobio,1989,135:2875-2883.[28]vandeWalleMV.,andShiloachJ.,BiotechnolBioeng,1998,57(1):71-78.[29]BauerKA.,Ben-BassatA.,DawsonM.etal.,ApplEnvironMicrobio,1990,56:1296-1302.[30]KlemanGL.,andStrohlWR.,ApplEnviroMicrobio,1994,60(11):3952-3958.[31]RiesenbergD.,SchulzV.,KnorreMA.,JBiotechnol,1991,20(1):17-28.[32]OhtaK.,ShibuiT.,MorimotoY.etal.,JFermentBioeng,1993,75(2):155-157.[33]LeeCW.,GuMB.,andChangHN.,EnzymeMicrobioTech-nol,1989,11:49-54.[34]SeoDJ.,ChungBH.,HwangYB.etal.,JFermentBioeng,1992,74:196-198.[35]ShimizuM.,HjimaS.,andKobayashiT.,JFermentBioeng,1992,74(3):163-168.[36],,,,1998,28(2):20-25.[37],,,,1993,19(2):153-158.ProgressStudiesofHighCell-densityCultureofRecombinantEscherichiacoliLiMin,ChenChangqing(ShanghaiResearchCenterofBiotechnology,ChineseAcademyofSciences,Shanghai200233)AbstractHighcell-densityfermentationofrecombinantE.coliwasanefficientapproachtoproveproduc-tionyieldandahotpointinfermentationengineering.Thisarticlediscussedseveralfactorsinfluencingthepro-ductivityincludinghost,medium,cultivationconditions,feedingstrategies