大腸桿菌分類

大腸桿菌分類Alistedgenenamemeansthatgenecarriesalossoffunctionmutation,a△precedingagenenamemeansthegeneisdeleted.Ifageneisnotlisted,itisnotknowntobemutated.ProphagespresentinwtK-12strains(F,入，e14,rac)arelistedonlyifabsent.E.coliBstrainsarenaturallylon-anddcm-.F-=DoesnotcarrytheFplasmidF+=CarriestheFplasmid.ThecellisabletomatewithF-throughconjugation.F'[]=CarriesanFplasmidthathashostchromosomalgenesonitfromapreviousrecombinationevent.ThiscellcanalsomatewithF-throughconjugation.ChromosomalgenescarriedintheFplasmidarelistedinbrackets.rB/K+/-=The(B/K)definesthestrainlineage.The+/-indicateswhetherthestrainhasorhasn'tgottherestrictionsystem.mB/K+/-=The(B/K)definesthestrainlineage.The+/-indicateswhetherthestrainhasorhasn'tgotthemodification(methylation)system.hsdS=Bothrestrictionandmethylationofcertainsequencesisdeletedfromthestrain.IfyoutransformDNAfromsuchastrainintoawildtypestrain,itwillbedegraded.hsdB=ForefficienttransformationofclonedunmethylatedDNAfromPCRamplificationsINV()chromosomalinversionbetweenlocationsindicatedahpC=mutationtoalkylhydroperoxidereductaseconferringdisulfidereductaseactivityara-14=cannotmetabolizearabinosearaD=mutationinL-ribulose-phosphate4-epimeraseblocksarabinosemetabolismcycA=mutationinalaninetransporter;cannotusealanineasacarbonsourcedapD=mutationinsuccinyldiaminopimelateaminotransferaseleadstosuccinateor(lysine+methionine)requirementA(=chromosomaldeletionofgenesbetweenthelistedgenes(mayincludeunlistedgenes!)dam=adeninemethylationatGATCsequencesexist;highrecombinationefficiency;DNArepairturnedondcm=cytosinemethylationatsecondCofCCWGGsitesexist.dam&dcmarethedefaultpropertiesandalwayselided,whiledam-ordcm-shouldbedeclareexplicitlyd￡QB=regulatorygenethatallowsconstitutiveexpressionofdeoxyribosesynthesisgenes;permitsuptakeoflargeplasmids.SeeHanahanD,USPatent4,851,348.***Thishasbeencalledintoquestion,astheDH10BgenomesequencerevealedthatitisdeoR+.SeeDurfee08,PMID18245285.dnaJ=oneofthechaparoninsinactivated;stabilizessomemutantproteinsdull=dUTPaseactivityabolished,leadingtoincreaseddUTPconcentrations,allowinguracilinsteadofthymineincorporationinDNA.StableUincorporationrequiresunggenemutationaswell.endAI=ForcleanerpreparationsofDNAandbetterresultsindownstreamapplicationsduetotheeliminationofnon-specificdigestionbyEndonucleaseI(e14)=excisableprophagelikeelementcontainingmcrAgene;presentinK-12butmissinginmanyotherstrainsgalE=mutationsareassociatedwithhighcompetence,increasedresistancetophageP1infection,and2-deoxygalactoseresistance.galEmutationsblocktheproductionofUDP-galactose,resultingintruncationofLPSglycanstotheminimal,"innercore".TheexceptionalcompetenceofDH10B/TOP10isthoughttobearesultofareducedinterferencefromLPSinthebindingand/oruptakeoftransformingDNA.galE15isapointmutationresultinginaSer123->Pheconversionneartheenzyme'sactivesite.SeevanDie,etal.PMID6373734,Hanahan,etal.PMID1943786,andEcoSalISBN1555811647.--Dcekiert16:56,23January2008(CST)galk=mutantscannotmetabolizegalactoseandareresistantto2-deoxygalactose.galK16isanIS2insertion~170bpdownstreamofthegalKstartcodon.SeeEcoSalISBN1555811647.--Dcekiert16:56,23January2008(CST)galU=mutantscannotmetabolizegalactosegor=mutationinglutathionereductase;enhancesdisulphidebondformationglnV=suppressionofamber(UAG)stopcodonsbyinsertionofglutamine;requiredforsomephagegrowthgyrA96=mutationinDNAgyrase;conveysnalidixicacidresistancegyrA462=mutationinDNAgyrase;conveysresistancetoccdBcolicingeneproducthflA150=proteasemutationstabilizingphagecIIprotein;highfrequencyoflysogenizationby入-A(lac)X74DeletionoftheentirelacoperonaswellassomeflankingDNA(completedeletionisAcod-mhpF;seeMol.Micro.,6:1335,andJ.Bact.,179:2573)lacIqorlacIQ=overproductionofthelacrepressorprotein;-35siteinpromoterupstreamoflacIismutatedfromGCGCAAtoGTGCAAlacIQ1=overproductionofthelacrepressorprotein;containsa15bpdeletiontocreateoptimal-35siteinpromoterupstreamoflacIlacY=deficientinlactosetransport;deletionoflactosepermease(Mprotein)lacZAM15=partialdeletionofthelacZgenethatallowsacomplementationofthep-galactosidasegene;requiredforblue/whiteselectiononXGalplates.DeletestheaminoportionoflacZ(aa11-41).leuB=requiresleucineAlon=deletionofthelonproteasemalA=cannotmetabolizemaltosemcrA=MutationeliminatingrestrictionofDNAmethylatedatthesequenceCmCGG(possiblymCG).Carriedonthee14prophage(q.v.)mcrB=MutationeliminatingrestrictionofDNAmethylatedatthesequenceRmCmetB=requiresmethioninemetC=requiresmethioninemrr=MutationeliminatingrestrictionofDNAmethylatedatthesequenceCmAGorGmACmtlA=cannotmetabilizemannitol(Mu)=Muprophagepresent.MuOmeansthephageisdefective.mutS-mutationinhibitsDNArepairofmismatchesinunmethylatednewlysynthesizedstrandsnupG=sameasdeoRompT=mutationinoutermembraneproteinproteaseVII,reducingproteolysisofexpressedproteins(P1)=CellcarriesaP1prophage.CellsexpresstheP1restrictionsystem.(P2)CellcarriesaP2prophage.AllowsselectionagainstRed+Gam+入仲80)Cellcarriesthelambdoidprophage^80.AdefectiveversionofthisphagecarryinglacZM15deletion(aswellaswild-typelacI,lacYA,andflankingsequences)ispresentinsomestrains.The^80attachmentsiteisjustadjacenttotonB.pLysS=containspLysSplasmidcarryingchloramphenicolresistanceandphageT7lysozyme,effectiveatattenuatingactivityofT7RNApolymerase,forbetterinhibitionofexpressionundernon-inducedconditions.TA/B=requiresprolinerecA1=ForreducedoccurrenceofunwantedrecombinationinclonedDNA;cellsUVsensitive,deficientinDNArepairrecA13=asforrecA1,butinsertslessstable.recBCD=ExonucleaseV;mutationinRecBorRecCreducesgeneralrecombinationbyafactorof100;impairedDNArepair;UVsensitive,easierpropagationofinvertedrepeatsrecJExonucleaseinvolvedinalternaterecombinationrelA=relaxedphenotype;permitsRNAsynthesisinabsenceofproteinsynthesisrha=blockedrhamosemetabolismrnc=encodesRnaselll(rnc-14isacommonnullmutant)rne=encodesRnaseE(rne-3071isacommontemperaturesensitivemutant)rpsL=mutationinribosomalproteinS12conveyingstreptomycinresistance;alsocalledstrAsbcBC=ExoIactivityabolished;usuallypresentinrecBCstrains;recombinationproficient,stableinvertedrepeatssr1=cannotmetabolizesorbitol-supE=glnVsupF=tyrTthi=requiresthiaminethyA=requiresthymidineTn10=transposonnormallycarryingTetracyclineresistanceTn5=transposonnormallycarryingKanamycinresistancetonA=MutationinoutermembraneproteinconveyingresistancetophageT1andphageT5traD=Mutationeliminatingtransferfactor;preventstransferofFplasmidtrxB=mutationinthioredoxinreductase;enhancesdisulphidebondformationinthecytoplasmtsx=outermembraneproteinmutationconveyingresistancetophageT6andcolicinKtyrT=suppressionofamber(UAG)stopcodonsbyinsertionoftyrosine;neededforsomephageinfectionsuchasAgt11.ung1=allowsuraciltoexistinplasmidDNAxyl-5=blockedxylosemetabolismSmR=StreptomycinresistanceMethylationIssuesinE.coliTypeImethylationsystems:E.coliK-12restrictsDNAwhichisnotprotectedbyadeninemethylationatsitesAA*C[N6]GTGCorGCA*C[N6]GTT,encodedbythehsdRMSgenes(EcoKI).Deletionsinthesegenesremoveseithertherestrictionormethylationorbothofthesefunctions.E.coliBderivativestrainscontainanhsdRMSsystem(EcoBI)restrictingandprotectiingthesequenceTGA*[N8]TGCTorAGCA*[N8]TCA.ThemcrAgene(carriedonthee14prophage)restrictsDNAwhichismethylatedinCmCWGGormCGsequences(methylationbythedcmgeneproduct).ThemcrBCgenesrestrictRmCsequences.ThemrrgeneproductrestrictsadeninemethylatedsequencesatCAGorGACsites.E.colimethylatestheadenineinGATC(andthecorrespondingAontheoppositestrand)withthedamgeneproduct.M.EcoKIImethylatesthefirstAatthepalindromicsiteATGCAT(aswellasthecorrespondingAontheoppositestrand),see(KossykhVG(2004)J.Bact186:2061-2067PMID15028690)Notethatthisarticlehasbeenretracted;theretractionappearstocenterontextualplagarism,notexperimentalresults.ThehomologytoAvaIIIisreal.IthinkIbelieveit.tk20:28,9December2005(EST).RichRobertsreports:"Wehavetriedourselvestodetectactivitywiththisgeneproductandcannotdetectanymethyltransferaseactivity.InourcaseweusedantibodiesabletodetectN6-methyladenineorN4methylcytosineinDNA.Theoneswehaveareverysensitiveandshouldhavebeenabletodetect5methylgroupsinthewholeE.colichromosome.Nothingwasdetectedinanoverexpressingstrain."ForadditionalinformationseeE.colirestriction-modificationsystemandtheNEBtechnicalinformationonmethylationCommonlyusedstrainsAG1endAIrecAIgyrA96thi-1relAIglnV44hsdR17(rKmK+)AB1157thr-1,araC14,leuB6(Am),A(gpt-proA)62,lacY1,tsx-33,qsr'-0,glnV44(AS),galK2(Oc),LAM-,Rac-0,hisG4(Oc),rfbC1,mgl-51,rpoS396(Am),rpsL31(strR),kdgK51,xylA5,mtl-1,argE3(Oc),thi-1BachmannBJ:DerivationandgenotypesofsomemutantderivativesofEscherichiacoliK-12.EscherichiacoliandSalmonellatyphimurium.CellularandMolecularBiology(Editedby:FCNeidhardtJLIngrahamKBLowBMagasanikMSchaechterHEUmbarger).Washington,D.C.,AmericanSocietyforMicrobiology1987,2:1190-1219.SeeCGSC#1157BL21E.coliBF-dcmompThsdS(rB-mB-)gal[malB+]K12(入S)The"malBregion"wastransducedinfromtheK-12strainW3110tomakethestrainMal+AS.SeeStudieretal.(2009)J.Mol.Biol.394(4),653foradiscussionoftheextentofthetransfer.StratageneE.coliGenotypeStrainsBL21(AI)F-ompTgaldcmlonhsdSB(rB-mB)araB::T7RNAP-tetAanE.coliBstraincarryingtheT7RNApolymerasegeneinthearaBlocusofthearaBADoperonq.TransformedplasmidscontainingT7promoterdrivenexpressionarerepresseduntilL-arabinoseinductionofT7RNApolymerase.DerivedfromBL21.Seetheproductpageformoreinformation.BL21(DE3)F-ompTgaldcmlonhsdSB(rB-mB-)A(DE3[lacIlacUV5-T7gene1ind1sam7nin5])anE.coliBstrainwithDE3,aAprophagecarryingtheT7RNApolymerasegeneandlacIqTransformedplasmidscontainingT7promoterdrivenexpressionarerepresseduntilIPTGinductionofT7RNApolymerasefromalacpromoter.DerivedfromB834(Wood,1966)bytransducingtoMet+.SeetheoriginalStudierpaperorthesummaryinMethodsinEnzymologyformoredetails.Wholegenomesequenceavailable[1]BL21(DE3)pLysSF-ompTgaldcmlonhsdSB(rB-mB-)A(DE3)pLysS(cmR)pLysSplasmidchloramphenicolresistant;growwithchloramphenicoltoretainplasmidChloramphenicolresistantThepLysSplasmidencodesT7phagelysozyme,aninhibitorforT7polymerasewhichreducesandalmosteliminatesexpressionfromtransformedT7promotercontainingplasmidswhennotinduced.seeMoffatt87fordetailsofpLysSandpLysEplasmidsBNN93F-tonA21thi-1thr-1leuB6lacY1glnV44rfbC1fhuA1mcrBe14-(mcrA-)hsdR(rK-mK+)A-SomeC600strainsarereallyBNN93BNN97-BNN93(入gt11)?A入gt11lysogenproducingphageat42CBW26434,CGSCStrain#7658△(araD-araB)567,△(lacA-lacZ)514(::kan),lacIp-4000(lacIq),A-,rpoS396(Am)?,rph-1,△(rhaD-rhaB)568,hsdR514ThisinformationisfromaprintoutsentbytheE.coliGeneticStockCenterwiththestrain.B.L.Wannerstrainrph-1isa1bpdeletionthatresultsinaframeshiftoverlast15codonsandhasapolareffectonpyrEleadingtosuboptimalpyrimidinelevelsonminimalmedium.(Jensen1993JBact.175:3401)△(araD-araB)567wasformerlycalledAaraBADaH33byDatsenkoandWannerAm=amber(UAG)mutationReference:DatsenkoandWanner,2000,PNAS,97:6640NOTE:ThispromoterdrivingtheexpressionoflacIwassequencedinthisstrainusingaprimerinmhpR(upstreamoflacI)andaprimerintheoppositeorientationinlacI.Thelacpromoterwasfoundtobeidenticaltowildtype.Thus,the-35sequencewasGCGCAAnotGTGCAAasexpectedwithlacIq.Thereforethisstrain(oratleasttheversionobtainedfromtheE.coliGeneticStockCenter)doesNOTappeartobelacIq.AccordingtoBarryWanner,thisisanunexpectedresult.-Reshma13:19,5May2005(EDT)"WehavenowconfirmedthatBW25113,BW25141,andBW26434arealllacI+,andnotlacIq.WethankyouforalertingustotheerrorwithrespecttoBW26434.Apparently,thelacIregionwasrestoredtowild-typeinapredecessorofBW25113."(fromBarryWannerNovember18,2005)ThegenotypehasbeencorrectedattheCGSCC600F-tonA21thi-1thr-1leuB6lacY1glnV44rfbC1fhuA1A-Therearestrainscirculatingwithbothe14+(mcrA+)ande14-(mcrA-)GeneralpurposehostSeeCGSC#3004References:Appleyard,R.K.(1954)Genetics39,440;Hanahan,D.(1983)J.Mol.Biol.166,577.C600hflA150(Y1073,BNN102)F-thi-1thr-1leuB6lacY1tonA21glnV44入-hflA150(chr::Tn10)hostforrepressingplaquesof入gt10whenestablishingcDNAlibrariesReferenceYoungR.A.andDavis,R.(1983)Proc.Natl.Acad.Sci.USA80,1194.TetracyclineresistancefromtheTn10insertionCSH50F-入-ara△(lac-pro)rpsLthifimE::IS1SeeCGSC#8085References:Miller,J.H.1972.Expts.inMolec.Genetics,CSH0:14-0;Blomfeldetal.,J.Bact.173:5298-5307,1991.D1210HB101lacIqlacY+DB3.1F-gyrA462endA1glnV44F(sr1-recA)mcrBmrrhsdS20(rB-,mB-)ara14galK2lacY1proA2rpsL20(Smr)xyl5△leumtl1usefulforpropagatingplasmidscontainingtheccdBoperon.gyrA462enablesccdBcontainingplasmidpropagationstreptomycinresistantappearstoNOTcontainlacI(basedonacolonyPCR)--AustinChe16:16,18June2007(EDT)<biblio>Bernard-JMolBiol-1992pmid=1324324Miki-JMolBiol-1992pmid=1316444</biblio>DH1endA1recA1gyrA96thi-1glnV44relA1hsdR17(rKmK+)A-parentofDH5aAnHoffman-Berling1100strainderivative(Meselson68)moreefficientattransforminglarge(40-60Kb)plasmidsnalidixicacidresistantReference:MeselsonM.andYuanR.(1968)Nature217:1110PMID4868368.DH5aF-endAIglnV44thi-1recAIrelAIgyrA96deoRnupG中80d/acZAM15L(lacZYA-argF)U169,hsdR17(rKmK+),A-AnHoffman-Berling1100strainderivative(Meselson68)PromegaalsolistsphoAnalidixicacidresistantReferences:FOCUS(1986)8:2,9.Hanahan,D.(1985)inDNACloning:APracticalApproach(Glover,D.M.,ed.),Vol.1,p.109,IRLPress,McLean,Virginia.Grant,S.G.N.etal.(1990)Proc.Natl.Acad.Sci.USA87:4645-4649PMID2162051.MeselsonM.andYuanR.(1968)Nature217:1110PMID4868368.DH10B(Invitrogen)F-endAIrecAIgalE15galK16nupGrpsLAlacX74080lacZAM15araD139△(ara,leu)7697mcrA△(mrr-hsdRMS-mcrBC)A-suitableforcloningmethylatedcytosineoradeninecontainingDNAanMC1061derivative(Casadaban80).PreparecellsforchemicaltransformationwithCCMB80bufferblue/whiteselectionWhileDH10BhasbeenclassicallyreportedtobegalUgalK,thepreliminarygenomesequenceforDH10BindicatesthatDH10B(andbytheirlineagealsoTOP10andanyotherMC1061derivatives)isactuallygalEgalKgalU+.Dcekiert16:37,23January2008(CST)GenomesequenceindicatesthatDH10BisactuallydeoR+.PresumablyTOP10andMC1061arealsodeoR+.StreptomycinresistantReferences:Casdaban,M.andCohen,S.(1980)JMolBiol138:179PMID6997493.Grant,S.G.N.etal.(1990)Proc.Natl.Acad.Sci.USA87:4645-4649PMID2162051.E.coliGeneticStockCenter,MC1061Record.DH10BGenomeSequencingProject,BaylorCollegeofMedicineCompletesequenceisavailable,seeDurfee08,PMID18245285.DH12S(Invitrogen)mcrA△(mrr-hsdRMS-mcrBC)^80dlacZAM15AlacX74recAIdeoRA(ara,leu)7697araD139galUgalKrpsLF'[proAB+lacIqZAM15]hostforphagemidandM13vectorsusefulforgeneratinggenomiclibrariescontainingmethylatedcytosineoradenineresiduesstreptomycinresistantReferences:Lin,J.J.,Smith,M.,Jessee,J.,andBloom,F.(1991)FOCUS13,96.;Lin,J.J.,Smith,M.,Jessee,J.,andBloom,F.(1992)BioTechniques12,718.DM1(Invitrogen)F-dam-13::Tn9(CmR)dcm-mcrBhsdR-M+gal1gal2ara-lac-thr-leu-tonRtsxRSu0HostforpBR322andothernon-pUC19plasmids;usefulforgeneratingplasmidsthatcanbecleavedwithdamanddcmsensitiveenzymesChloramphenicolresistantPromegalistsasF'notF-Reference:Lorow-MurrayDandBloomF(1991)Focus13:20E.cloni(r)5alpha(Lucigen)fhuA24(argF-lacZ)U169phoAglnV44(^80^(lacZ)M15gyrA96recAIrelAIendAIthi-1hsdR17Commoncloningstrain.E.cloni(r)10G(Lucigen)F-mcrA△(mrr-hsdRMS-mcrBC)endA1recAI080dlacZAM15AlacX74araD139△(ara,/eu)7697galUgalKrpsLnupGX-tonACommoncloningstrain.ResistanttophageT1.E.cloni(r)10GF'(Lucigen)[F'proA+B+lacIqZAM15::Tn10(TetR)]/mcrA△(mrr-hsdRMS-mcrBC)endA1recA1080dlacZAM15△lacX74araD139△(ara,leu)7697galUgalKrpsLnupGXtonAStrainforcloningandsingle-strandDNAproduction.E.coliK12ER2738(NEB)F'proA+B+laclq△(lacZ)M15zzf::Tn10(TetR)/fhuA2glnV△(lac-proAB)thi-1△(hsdS-mcrB)5PhagepropagationstrainAlsoavailablefromLucigenCorporation.ER2566(NEB)F-入-fhuA2[lon]ompTlacZ::T7gene1galsulA11F(mcrC-mrr)114::IS10R(mcr-73::miniTn10-TetS)2R(zgb-210::Tn10)(TetS)endA1[dcm]HoststrainfortheexpressionofatargetgeneclonedinthepTYBvectors.CarryachromosomalcopyoftheT7RNApolymerasegeneinsertedintolacZgeneandthusunderthecontrolofthelacpromoter.IntheabsenceofIPTGinductionexpressionofT7RNApolymeraseissuppressedbythebindingoflacIrepressortothelacpromoter.DeficientinbothlonandompTproteases.ER2267(NEB)F'proA+B+lacIqF(lacZ)M15zzf::mini-Tn10(KanR)/F(argF-lacZ)U169glnV44e14-(McrA-)rfbD1?recA1relA1?endA1spoT1?thi-1△(mcrC-mrr)114::IS10CommonlyusedfortiteringM13phagebecauseofthestrain'sF'plasmid,whichcarriesKanR,anditsslowgrowth,whichpromoteseasyvisualizationofplaques.HB101F-mcrBmrrhsdS20(rB-mB-)recA13leuB6ara-14proA2lacY1galK2xyl-5mtl-1rpsL20(SmR)glnV44A-Pleasenotethatdifferentsourceshavedifferentgenotypessotreatthisinformationwithcaution.FromaGIBCOBRLlistofcompetentcells.HybridofE.coliK12andE.coliB(but98%KstrainAB266accordingtoSmithetal.)HostforpBR322andmanyplasmidsSigmaliststhedeletion△(gpt,proA).Checkthis.PromegadoesnotlistF-,mcrB,ormrrStreptomycinresistantReferences:-Boyer,H.W.andRoulland-Dussoix,D.(1969)J.Mol.Biol.41,459.Smith,M.,Lorow,D.,andJessee,J.(1989)FOCUS11,56-pdfversionfromInvitrogenLacksSandGreenbergJR(1977)JMolBiol114:153.HMS174(DE3)F-recA1hsdR(rK12-mK12+)(DE3)(RifR)HMS174strainsprovidetherecAmutationinaK-12background.LikeBLR,thesestrainsmaystabilizecertaintargetgeneswhoseproductsmaycausethelossoftheDE3prophage.DE3indicatesthatthehostisalysogenoflDE3,andthereforecarriesachromosomalcopyoftheT7RNApolymerasegeneundercontrolofthelacUV5promoter.SuchstrainsaresuitableforproductionofproteinfromtargetgenesclonedinpETvectorsbyinductionwithIPTG.High-Control(tm)BL21(DE3)(Lucigen)F-ompTgaldcmhsdSB(rB-mB-)(DE3)/Mini-F/ac/q1(Gentr)TheHI-ControlBL21(DE3)cellscontainasingle-copyBACplasmidharboringaspeciallyengineeredversionofthe/ac/q1repressorallele.The/ac/q1alleleexpresses~170-foldmorelacrepressorproteinthanthewild-type/ac/gene.TheincreasedpooloflacrepressorinHI-ControlBL21(DE3)cellsmaintainstightcontrolovertheexpressionofT7RNApolymerasefromthelacUV5promoter,reducingleakyexpressionofgenesclonedunderaT7promoter.anE.co/iBstrainwithDE3,a入prophagecarryingtheT7RNApolymerasegeneandlacIqTransformedplasmidscontainingT7promoterdrivenexpressionarerepresseduntilIPTGinductionofT7RNApolymerasefromalacpromoter.High-Control(tm)10G(Lucigen)F-mcrA△(mrr-hsdRMS-mcrBC)endA1recA108Gd/acZAM15△/acX74araD139△(ara,/eu)7697ga/Uga/KrpsLnupGA-tonA/Mini-F/ac/q1(Gentr)TheHI-Control10Gcellscontainasingle-copyBACplasmidharboringaspeciallyengineeredversionofthe/ac/q1repressorallele.The/ac/q1alleleexpresses~170-foldmorelacrepressorproteinthanthewild-type/ac/gene.ForstablecloningofT7proteinexpressionplasmids.ResistanttophageT1.IJ1126E.coliK-12recB21recC22sbcA5endAgalthiSu+A(mcrC-mrr)102::Tn10SeeEndy:IJ1126IJ1127IJ1126lacUV5lacZ::T7gene1-KnrSeeEndy:IJ1127JM83rpsLara△(lac-proAB)080dlacZAM15Sigmaliststhi.Checkthis.streptomycinresistantJM101glnV44thi-1A(lac-proAB)F'[lacIqZAM15traD36proAB+]hostforM13mpvectorsrecA+,rK+originalblue/whitecloningstrainhasallwtrestrictionsystemsReferences:Messing,J.etal.(1981)NucleicAcidsRes.9,309;Yanisch-Perron,C.,Vieira,J.,andMessing,J.(1985)Gene33,103.JM103endA1glnV44sbcBCrpsLthi-1△(lac-proAB)F'[traD36proAB+lacIqlacZAM15]streptomycinresistantReferences:Hanahan,D.(1983)J.Mol.Biol.166:557-80.NEBsaysthisstrainencodesaprophageencodedEcoP1endonuclease.Sigmalists(P1)(r-m+rP1+mP1+)JM105endA1glnV44sbcB15rpsLthi-1△(lac-proAB)[F'traD36proAB+lacIqlacZAM15]hsdR4(rK-mK+)SigmalistssbcCstreptomycinresistantReferences:Yanisch-Perron,C.,Vieira,J.,andMessing,J.(1985)Gene33,103.JM106endA1glnV44thi-1relA1gyrA96△(lac-proAB)F-hsdR17(rK-mK+)References:Yanisch-Perron,C.,Vieira,J.,andMessing,J.(1985)Gene33,103.JM107endA1glnV44thi-1relA1gyrA96△(lac-proAB)[F'traD36proAB+lacIqlacZAM15]hsdR17(RKmK+)入-hostforM13mpvectorsrecA+,rK+Sigmalistse14-(McrA-)nalidixicacidresistantReferences:Yanisch-Perron,C.,Vieira,J.,andMessing,J.(1985)Gene33,103.JM108endA1recA1gyrA96thi-1relAIglnV44A(lac-proAB)hsdR17(rK-mK+)nalidixicacidresistantdeficientinexpressionofthelonproteaseduetoIS186transposoninsertion--J_Mairhofer18:59,24March2010(CET)<biblio>1.Referencepmid=20138928</biblio>JM109endA1glnV44thi-1relA1gyrA96recA1mcrB+△(lac-proAB)e14-[F'traD36proAB+lacIqlacZ^M15]hsdR17(rKmK+)-FromNEBPartlyrestriction-deficient;goodstrainforcloningrepetitiveDNA(RecA-).SuppressesmanyambermutationswhenglutamineisacceptablebutnottheS100orS7mutationsof入，e.g.,入gt11.CanalsobeusedforM13cloning/sequencingandblue/whitescreening.Sigmalistse14-nalidixicacidresistantdeficientinexpressionofthelonproteaseduetoIS186transposoninsertion--J_Mairhofer18:59,24March2010(CET)FromC.Yanisch-Perron,J.Vieira,andJ.Messing.ImprovedM13phagecloningvectorsandhoststrains:nucleotidesequencesoftheM13mp18andpUC19vectors.Gene,33(1):103-19,1985.SomeinformationfromMaryBerlynattheE.coliGeneticStockCenter:OneofthereasonstheoriginalcuratorofthiscollectiondidnotaccessiontheJM109,JM103,etc.strainswasbecauseshefounditimpossibletobesureofthederivationandthereforethedetailsofthegenotype.ButIthinkit'ssafetoassumethattheF'inthisstrainisderivedfromorsimilartoF128whichextendsfromtheproBAregionthroughthelacoperon.ItthuscarriesthewildtypegenesforalllociinthatregionexceptthoseindicatedasmutantforthegenotypeoftheF'.SoitcarriesthelacZ(alpha-complementation)deletionlacZ58(M150andthelacImutationlacIq,butithasthelacY+genealsoontheF-prime.Onthechromosomeitlacksallthelacoperongenes.NOTE:ThepromoterdrivingtheexpressionoflacIwassequencedinthisstrainusingaprimerinmhpR(upstreamoflacI)andaprimerintheoppositeorientationinlacI.Thelacpromoterwasfoundtobeidenticaltowildtype.Thus,the-35sequencewasGCGCAAnotGTGCAAasexpectedwithlacIQ.Thereforethisstrain(oratleasttheversionwehave)doesNOTappeartobelacIQunlessthereisanothercopyoflacIelsewhere.ThisresultissomewhatconfirmedbythefactthatalacIregulatedpromoterdrivingexpressionofYFPonamediumcopyvectordoesnotrepresscompletely.-Reshma13:48,5May2005(EDT)JM109(DE3)JM109+A(DE3)DE3prophagecarryingT7polymeraseexpressioncassetteSamecassetteasBL21(DE3)carryingalacinducibleT7RNApolymeraseandlacIqnalidixicacidresistantJM110rpsLthrleuthilacYgalKgalTaratonAtsxdamdcmglnV44A(lac-proAB)e14-[F'traD36proAB+lacIqlacZAM15]hsdR17(rKmK+)SigmafailstolisttonAtsxe14fhuAhsdR17(e14-)statusuncertainstreptomycinresistantJM2.300lacI22,LAM-,e14-,rpsL135(strR),malT1(LamR),xyl-7,mtl-1,thi-1Somefolkshavebeenusingthisstrain(i.e.,Elowitz,Gardner)andittookmetoolongtofindtheCGSC#.ThisstrainisnolongeravailablefromtheCGSCLE392glnV44supF58(lacY1orAlacZY)galK2galT22metBItrpR55hsdR514(rK-mK+)SigmalistsF-e14-MachlArecA1398endA1tonA中80AlacM15AlacX74hsdR(rKmK+)FromInvitrogenDoublingtimeapprox.50minandsupposedlyfastestgrowingchemicallycompetentcloningstrainavailableMachlcellsarederivativesofE.coliWstrains(ATCC9637,S.A.Waksman),ratherthanE.coliK-12.ThismayhaveimplicationsforBL-1statusforsomefacilities(apparentlynotforMIT).SeeBloom04patentfordetailsontheconstructionandpropertiesofthisstrain.MC1061F-F(ara-leu)7697[araD139]B/rF(codB-lacI)3galK16galE15入-e14-mcrA0relA1rpsL150(strR)spoT1mcrB1hsdR2(r-m+)StreptomycinresistantThethr-leuregionwastransducedfromanE.coliB/rstrain(SB3118)inearlystepsofstrainconstruction.ParentofDH10B/TOP10andderivedstrainsReferences:E.coliGeneticStockCenter,MC1061RecordCasdaban,M.andCohen,S.(1980)JMolBiol138:179PMID6997493.CompleteDH10Bsequenceisavailable,seeDurfee08,PMID18245285.MC4100F-[araD139]B/r△(argF-lac)169*&lambda-e14-flhD5301△(fruK-yeiR)725(fruA25井relAIrpsL150(strR)rbsR22△(fimB-fimE)632(::IS1)deoC1Thethr-leuregionwastransducedfromanE.coliB/rstrain(SB3118)inearlystepsofstrainconstruction.ThispapercomparesMC4100toMG1655anddescribesthesignificantdeletions.*Thepaperreferencedaboveshowedthatthisdeletionwaslargerthanpreviouslyknown.ThedeletionnowcoversykfD-b0350.*ThefruA25alleleisattributedtothedeletionoffruK-yeiR.ThismeansfruAispresentbutitspromoterhasbeendeleted.Thepaperalsoshowsthatthee14elementisdeletedinMC4100.OneofthegenesremovedbythisdeletionismcrA,whichencodesanenzymethatrestrictsDNAcontainingmethylcytosine.However,otherE.colK-12restriction/modificationsystemsarestillpresentinMC4100.MC4100stillencodestheMcrBC5-methylcytosine=specificrestrictionenzymeandtheHsdR/HsdS/HsdMtypeIrestriction-modificationcomplex.TablethreeofthepaperlistsallgenesbelievedtobedeletedinMC4100.Themethodsusedinthepapercandetectdeletionsbutnotlossoffunctionmutations.SeeCGSC#6152MG1655F-入-ilvG-rfb-50rph-1Thisisthe"wildtype"K-12strainwhichwassequenced,andshouldbeusedwhenPCRinggenesfromthesequencedgenome.Italsolooksveryhealthyunderthemicroscope--adramaticdifferencefrommostofthecloningstrains,whichappearsick.SeeCGSC#6300SeeATCC700926<biblio>1.Blattner-Science-1997pmid=9278503</biblio>Moreaccuratesequencecorrecting243errorsintheoriginalsequencingHoriuchi2006.NewGenbankaccessionnumberU00096.2OmniMAX2FromInvitrogen:"ThisstrainoverexpressestheLacrepressor(lacIqgene).Forblue/whitescreening,youwillneedtoaddIPTGtoinduceexpressionfromthelacpromoter.StrainisresistanttoT1bacteriophage."F{proAB+lacIqlacZAM15Tn10(TetR)△(ccdAB)}mcrA△(mrr-hsdRMS-mcrBC)^80(lacZ)AM15A(lacZYA-argF)U169endA1recA1supE44thi-1gyrA96relA1tonApanDOverExpress(tm)C41(DE3)(Lucigen)F-ompTgaldcmhsdSB(rB-mB-)(DE3)TheOverExpressstrainscontaingeneticmutationsphenotypicallyselectedforconferringtolerancetotoxicproteins.ThestrainC41(DE3)wasderivedfromBL21(DE3).Thisstrainhasatleastoneuncharacterizedmutation,whichpreventscelldeathassociatedwithexpressionofmanyrecombinanttoxicproteins.ThestrainC43(DE3)wasderivedfromC41(DE3)byselectingforresistancetoadifferenttoxicproteinandcanexpressadifferentsetoftoxicproteinstoC41(DE3).anE.coliBstrainwithDE3,a入prophagecarryingtheT7RNApolymerasegeneandlacIqTransformedplasmidscontainingT7promoterdrivenexpressionarerepresseduntilIPTGinductionofT7RNApolymerasefromalacpromoter.OverExpress(tm)C41(DE3)pLysS(Lucigen)F-ompTgaldcmhsdSB(rB-mB-)(DE3)pLysS(Cmr)TheOverExpressstrainscontaingeneticmutationsphenotypicallyselectedforconferringtolerancetotoxicproteins.ThestrainC41(DE3)wasderivedfromBL21(DE3).Thisstrainhasatleastoneuncharacterizedmutation,whichpreventscelldeathassociatedwithexpressionofmanyrecombinanttoxicproteins.ThestrainC43(DE3)wasderivedfromC41(DE3)byselectingforresistancetoadifferenttoxicproteinandcanexpressadifferentsetoftoxicproteinstoC41(DE3).anE.coliBstrainwithDE3,a入prophagecarryingtheT7RNApolymerasegeneandlacIqTransformedplasmidscontainingT7promoterdrivenexpressionarerepresseduntilIPTGinductionofT7RNApolymerasefromalacpromoter.ThepLysSplasmidencodesT7phagelysozyme,aninhibitorforT7polymerasewhichreducesandalmosteliminatesexpressionfromtransformedT7promotercontainingplasmidswhennotinduced.OverExpress(tm)C43(DE3)(Lucigen)F-ompTgaldcmhsdSB(rB-mB-)(DE3)TheOverExpressstrainscontaingeneticmutationsphenotypicallyselectedforconferringtolerancetotoxicproteins.ThestrainC41(DE3)wasderivedfromBL21(DE3).Thisstrainhasatleastoneuncharacterizedmutation,whichpreventscelldeathassociatedwithexpressionofmanyrecombinanttoxicproteins.ThestrainC43(DE3)wasderivedfromC41(DE3)byselectingforresistancetoadifferenttoxicproteinandcanexpressadifferentsetoftoxicproteinstoC41(DE3).anE.coliBstrainwithDE3,a入prophagecarryingtheT7RNApolymerasegeneandlacIqTransformedplasmidscontainingT7promoterdrivenexpressionarerepresseduntilIPTGinductionofT7RNApolymerasefromalacpromoter.OverExpress(tm)C43(DE3)pLysS(Lucigen)F-ompTgaldcmhsdSB(rB-mB-)(DE3)pLysS(Cmr)TheOverExpressstrainscontaingeneticmutationsphenotypicallyselectedforconferringtolerancetotoxicproteins.ThestrainC41(DE3)wasderivedfromBL21(DE3).Thisstrainhasatleastoneuncharacterizedmutation,whichpreventscelldeathassociatedwithexpressionofmanyrecombinanttoxicproteins.ThestrainC43(DE3)wasderivedfromC41(DE3)byselectingforresistancetoadifferenttoxicproteinandcanexpressadifferentsetoftoxicproteinstoC41(DE3).anE.coliBstrainwithDE3,a入prophagecar