Description:
TheWarmStartLAMPKit(DNA&RNA)isdesignedtoprovideasimple,one-stepsolutionforLoop-MediatedIsothermalAmplification(LAMP)ofDNAorRNA(RT-LAMP)targets.LAMPandRT-LAMParecommonlyusedisothermalamplificationtechniquesthatprovidesrapiddetectionofatargetnucleicacidusingLAMP-specificprimers(suppliedbytheuser)andastrand-displacingDNApolymerase.ThiskitissuppliedwiththeWarmStartLAMP2XMasterMix,whichcontainsablendofBst2.0WarmStartDNAPolymeraseandWarmStartRTxReverseTranscriptaseinanoptimizedLAMPbuffersolution.BothBst2.0WarmStartDNAPolymeraseandWarmStartRTxReverseTranscriptasehavebeenengineeredforimprovedperformanceinLAMPandRT-LAMPreactions.Afluorescentdyeisalsosuppliedtoenablereal-timefluorescencemeasurementofLAMP.TheWarmStartLAMPKitiscompatIBLewithmultipledetectionmethods,includingturbiditydetection,real-timefluorescencedetection(whenusedwithLAMPfluorescentdye)andend-pointvisualization.
NEB’sWarmStartLAMPKit(DNA&RNA)iscompatiblewithmultipledetectionmethods

TheNEBWarmStartLAMPKit(DNA&RNA)includesseparatefluorescentdyeforreal-timefluorescencemeasurement.Alternately,detectioncanbeaccomplishedbyturbiditydetectionorendpointvisualization.
NEB’sWarmStartLAMPKit(DNA&RNA)offersspeedandrobustsensitivity

ARNAtarget(HMBS2)wasamplifiedfromJurkattotalRNAusingtheWarmStartLAMPKitandOptigeneMasterMix(ISO-001).Reactionswereperformedat65°Cfor74minutesonareal-timeThermocycler(Bio-RadCFX96)intriplicate.Timetoresultissetasthetimeatwhichthefluorescencecrossedathresholdof10%ofmaximalfluorescence.NEB’sWarmStartLAMPKitresultedinfasterandmorerobustdetectionascomparedtotheOptiGeneMasterMix.
NEB’sWarmStartLAMPKit(DNA&RNA)providesfastLAMP/RT-LAMPresults

ExperimentsweredesignedtoamplifyaDNAtarget(BRCAb)andaRNAtarget(HMBS2)byLAMPorRT-LAMP,respectively.A25μlreactioncontaining10ngHeLaDNAor5ngJurkattotalRNA,1XMasterMix,1XLAMPprimersandfluorescentdyewasincubatedonaBio-RadCFX98instrumentat65°Cfor75minutes.Lucigen®OmniAmp®RNA&DNAKitandOptiGeneIsothermalMasterMix(ISO-001)wererunaccordingtomanufacturers’recommendations.NEBWarmStartLAMPKit(DNA&RNA)enablesspecificamplificationacrossmultiplespecies/targets
PRIMERSET | DNATEMPLATE |
---|
HeLa DNA | Lyme DNA | B.malayi DNA | E.coli DNA | C.elegans DNA |
---|
BRCAb(human) | + | | | | |
CFTR(human) | + | | | | |
LymO(Lyme) | | + | | | |
BmHhal(B.malayi) | | | + | | |
dnaE(E.coli) | | | | + | |
Lec10b(C.elegans) | | | | | + |
Lec6(C.elegans) | | | | | + |
Ineachindividualreaction,multiplegDNAtemplateswereaddedalongwithprimersspecifictoatargetinonlyonespecies.Reactionswereincubatedat65°Cfor74minutesonathermocycler(Bio-RadCFX96)intriplicate.Positiveresults(+)wereobservedonlyintheexpectedspecies/targetcombinations,demonstratinghighspecificityoftheWarmStartLAMPreactions.
KitComponents
Thefollowingreagentsaresuppliedwiththisproduct:
| Storeat(°C) | Concentration |
WarmStartLAMP2XMasterMix | -20 | |
LAMPFluorescentDye(50X) | -20 | |