Loop-Mediated Isothermal Amplification - NEB

Loop-mediated isothermal amplification (LAMP) uses 4-6 primers recognizing 6-8 distinct regions of target DNA. A strand-displacing DNA polymerase initiates ... Home Applications DNAAmplification,PCRandqPCR IsothermalAmplification Loop-MediatedIsothermalAmplification Loop-mediatedisothermalamplification(LAMP)uses4-6primersrecognizing6-8distinctregionsoftargetDNAforahighlyspecificamplificationreaction.Astrand-displacingDNApolymerase initiatessynthesisand2speciallydesignedprimersform“loop”structurestofacilitatesubsequentroundsofamplificationthroughextensionontheloopsandadditionalannealingofprimers.DNAproductsareverylong(>20kb)andformedfromnumerousrepeatsoftheshort(80–250bp)targetsequence,connectedwithsingle-strandedloopregionsinlongconcatamers.Theseproductsarenottypicallyappropriatefordownstreammanipulation,buttargetamplificationissoextensivethatnumerousmodesofdetectionarepossible.Real-timefluorescencedetectionusingintercalatorsorprobes,lateralflowandagarosegeldetectionarealldirectlycompatiblewithLAMPreactions.InstrumentationforLAMPtypicallyrequiresconsistentheatingtothedesiredreactiontemperatureand,whereneeded,real-timefluorescenceforquantitativemeasurements.OptimizedsettingsforrunningLAMPexperimentsonisothermalinstrumentssuchastheAxxinT8-ISOandT16-ISOcanbefound here. ReactionTemperature AmpliconSize DetectionMethod(s)  65°C  <250nt  Visual,Lateralflow,Gel,Turbidity   Loop-mediatedisothermalamplification(LAMP)uses4-6primersrecognizing6-8distinctregionsoftargetDNA.Astrand-displacingDNApolymeraseinitiatessynthesisand2oftheprimersformloopstructurestofacilitatesubsequentroundsofamplification. Inadditiontothemoretraditionalorcomplexdetectionmethods,LAMPissoprolificthattheproductsandbyproductsofthesereactionscanalsobevisualizedbyeye.Forexample,magnesiumpyrophosphateproducedduringthereactioncanbeobservedasawhiteprecipitateoraddedindicatorslikecalceinorhydroxynaphtholbluecanbeusedtosignalapositivereaction.Alternatively,usingthe WarmStart®2XColorimetricLAMPMasterMix developedbyNEBenablesastrongcolorchangefrompinktoyellowbasedonapHchangeduringthereaction. AnupdatedversionofthisproducthasbeenformulatedwithdUTPandUDGtobecompatiblewithcarryoverpreventionbetweenamplificationrounds–WarmStartColorimetricLAMP2XMasterMixwithUDG.ThecolorimetricdetectiontechnologyisakeycomponentoftheSARS-CoV-2RapidColorimetricLAMPAssayKit,whichcanbeusedintheanalysisofSARS-CoV-2,thenovelcoronavirusthatcausesCOVID-19. DesigningLAMPprimerscanbechallenging,butsoftwaretoolsgreatlyfacilitatethisprocess.Wesuggestusingthe NEBLAMPPrimerDesignTooltodesignLAMPprimers.AfterinputtingaDNAorRNAsequenceofinterest,theLAMPPrimerDesigntoolwillidentifysuitabletargetregionsandcreatetheouterF3/B3andloopinginnerFIP/BIPprimersinasinglestep.TheLoopF/LoopBprimers,thatacceleratetheLAMPreaction,arecreatedinasecondstepandarestronglyrecommendedforbestperformance. LAMPiswell-suitedforpoint-of-careandfielddiagnostics andLAMPassayshavebeendesignedforthedetectionofawiderangeofRNAandDNAtargetsfromallmannerofsampletypes.Examplesincludetestsfor: Rapiddetectionofcoronavirus(COVID-19)RNA  DirectdetectionofSARS-CoV-2RNAfromUniversalTransportMedium Filariasisinhumansandinsects Foodandwaterqualityin-fieldapplications DetectionoftheZikavirusinhumansamples TheLAMPreactionisrobustandtolerantofinhibitors,allowingforcrudesampleprepandminimalnucleicacidpurificationifdesired.WarmStart®RTx andBst2.0WarmStart weredevelopedforoptimalperformanceinLAMP/RT-LAMPandarecombinedinconvenientLAMPMasterMixes tosimplifyassaydesign.   ChooseType: FAQs Protocols ApplicationNotes Tools&Resources LegalInformation FAQs Protocols ApplicationNotes Tools&Resources LegalInformation FAQsforLoop-MediatedIsothermalAmplification HowcanIseetheproductsofaLAMPreaction? HowdoIuseTteUvrDHelicaseforreducingnon-templateamplificationinLAMPreactions? DoesNEBhaveamastermixforLAMPorRT-LAMPreactions? WhatisthedifferencebetweenBstDNAPolymerase,LargeFragment,Bst2.0,andBst3.0DNAPolymerase? HowdoIuseWarmStart®RTxinRT-LAMP? HowdoIuseAntarcticThermolabileUDGforcarryoverpreventioninLAMPreactions? ProtocolsforLoop-MediatedIsothermalAmplification Loop-mediatedIsothermalAmplification(LAMP) WarmStartLAMPKit(DNA&RNA)Protocol(E1700) WarmStartColorimetricLAMP2XMasterMixTypicalLAMPProtocol(M1800) ProtocolforLAMPReactionswithTteUvrDHelicase(#M1202) ProtocolforunwindingdoublestrandedDNAwithTteUvrDHelicase(#M1202) TypicalLAMPProtocol(M0275) TypicalLAMPProtocol(M0538) TypicalLAMPProtocol(M0374) TypicalRT-LAMPProtocol WarmStart®Multi-PurposeLAMP/RT-LAMP2XMasterMix(withUDG)Protocol(NEB#M1708) WarmStart®FluorescentLAMP/RT-LAMPKit(withUDG)Protocols(NEB#E1708) WarmStartColorimetricLAMP2XMasterMixwithUDGTypicalLAMPProtocol(NEB#M1804) SARS-CoV-2RapidColorimetricLAMPDetectionAssayProtocol(NEB#E2019) ApplicationNotesforLoop-MediatedIsothermalAmplification FacilitatingDetectionofSARS-CoV-2DirectlyfromPatientSamples:PrecursorStudieswithRT-qPCRandColorimetricRT-LAMPReagents OptimizedIntegrationofNewEnglandBiolabs®Loop-mediatedIsothermalAmplification(LAMP)ReagentswithAxxinISOInstruments Tools&Resources FeatureArticles MitigatingRiskandEnsuringConsistentSupplyChainThroughInternalSARS-CoV-2TestingwithRT-LAMP ThisfeaturearticledescribeshowNEBscientistsestablishedaCLIA-certifiedSARS-CoV-2onsitetestingprogramutilizingsalivaandcolorimetricloop-mediatedisothermalamplification(LAMP) Brochures IsothermalAmplificationTrifold WebTools NEBLAMPPrimerDesignTool Posters AmplificationReagentsforMolecularDiagnosticsApplications(2017) ColorimetricLAMP:VisualDetectionforSimpleDiagnostics(2017) Genomefilteringidentifiesspecies-specificDNAbiomarkersforMansonellaperstansandMansonellaozzardi,whichenabledifferentiationofthesecloselyrelatedspeciesandotherco-endemicfilarialparasites(2019) LegalInformation Thisproductiscoveredbyoneormorepatents,trademarksand/orcopyrightsownedorcontrolledbyNewEnglandBiolabs,Inc(NEB). WhileNEBdevelopsandvalidatesitsproductsforvariousapplications,theuseofthisproductmayrequirethebuyertoobtainadditionalthirdpartyintellectualpropertyrightsforcertainapplications. Formoreinformationaboutcommercialrights,pleaseemailusat [email protected]. Thisproductisintendedforresearchpurposesonly.Thisproductisnotintendedtobeusedfortherapeuticordiagnosticpurposesinhumansoranimals. Videos + NEB®LAMPPrimerDesignToolTutorial ThisvideoexplainshowtouseourLAMPPrimerDesignwebtool,whichmakesthecomplextaskofLAMPprimerdesignaseasyassubmittingyoursequence. + LoopMediatedIsothermalAmplification(LAMP)Tutorial Didyouknowthatthisisothermalamplificationmethodcanbeperformedinaslittleas5-10minuteswithlimitedresources?GetaquickoverviewonhowLAMPworksinthisanimation. + BehindthePaper:VisualDetectionofIsothermalNucleicAcidAmplificationUsingpH-sensitiveDyes OnelittleprotoncouldchangethewayscientistsdetectDNAamplificationinthefieldandpoint-of-caresettings.Nathansharesthedetailsofhisrecentpaper. + NEB®TVEp.23–ColorimetricLAMPinpoint-of-carediagnostics LearnthebasicsofColorimetricLAMP,andhowgroupslikeNEBandtheWorldMosquitoProgramareusingthisisothermalamplificationmethodinfieldtestsandpoint-of-carediagnostics. FeaturedProducts WarmStart®ColorimetricLAMP2XMasterMix(DNA&RNA) WarmStart®ColorimetricLAMP2XMasterMixwithUDG SARS-CoV-2RapidColorimetricLAMPAssayKit WarmStart® Multi-PurposeLAMP/RT-LAMP2XMasterMix(withUDG) WarmStart® FluorescentLAMP/RT-LAMPKit(withUDG) WarmStart®LAMPKit(DNA&RNA) Bst2.0DNAPolymerase WarmStart®RTxReverseTranscriptase Chooseyourcountry NorthAmerica Canada UnitedStates Europe France Germany UnitedKingdom Asia-Pacific Australia China Japan NewZealand Singapore Ifyoudon'tseeyourcountryabove,pleasevisitour internationalsite SessionExpired Youhavebeenidleformorethan20minutes,foryoursecurityyouhavebeenloggedout.Pleasesignbackintocontinueyoursession. SignIn InstitutionChanged YourprofilehasbeenmappedtoanInstitution,pleasesignbackforyourprofileupdatestobecompleted. SignIn SignintoyourNEBaccount Tosaveyourcartandviewpreviousorders,signintoyourNEBaccount.Addingproductstoyourcartwithoutbeingsignedinwillresultinalossofyourcartwhenyoudosigninorleavethesite. 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