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Epigentek/EpiNext High-Sensitivity DNA Library Preparation Kit (Illumina)/P-1053-24/24 reactions

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InputType:DNAResearchArea:NextGenSequencingTargetApplication:LibraryConstructionVesselFormat:Columns/Tubes100%Guarantee:6monthsProductOverviewTheEpiNext™ High-SensitivityDNALibraryPreparationKit(Illumina) isacompletesetofoptimizedreagentstoprepareaDNAlibraryfromverysmallamountofsamplesforuseinnext-generationsequencingapplications.ThiskitissuitableforpreparingaDNAlibraryusingsub-nanogramamountsofDNAinputfornextgenerationsequencingapplicationsusinganIlluminasequencer.TheseapplicationsincludegenomicDNA-seq,ChIP-seq,MeDIP/hMeDIP-seq,trADItionalbisulfite-seq,andtargetedre-sequencing.Theoptimizedprotocolandcomponentsofthekitallowbothnon-barcoded(singleplexed)andbarcoded(multiplexed)DNAlibrariestobeconstructedquicklywithreducedbias.Thekithasthefollowingadvantages:Highsensitivityandflexibility -Canbeusedforbothnon-barcoded(singleplexed)andbarcoded(multiplexed)DNAlibrarypreparation.TheamountofinputDNAcanbeaslowas0.2ngwitharangefrom0.2to100ng.VariousdsDNAcanbeused,whichincludeslimitedamountsoffragmenteddsDNAisolatedfromvarioustissueorcellsamples,dsDNAenrichedfromChIPreactions,anddsDNAenrichedfromMeDIP/hMeDIPreactionsorexoncapture. Fastandstreamlinedprocedure-TheprocedurefromfragmentedDNAtosizeselectionislessthan1hourand30minutes.Noclean-upisrequiredbetweeneachstepandallreactionstakeplaceinthesametube,therebysavingtimeandpreventinghandlingerrorsaswellaslossofvaluablesamples.Gel-freesizeselectionfurtherreducesthepreparationtime.Highlyconvenient-ThekitcontainsallrequiredcomponentsforeachstepofDNAlibrarypreparation,whicharesufficientforendpolishing,ligation,clean-up,sizeselection,andlibraryamplification,therebyallowingthelibrarypreparationtobestreamlinedwiththemostreliableandconsistentresults. Minimizedbias-UltraHiFiamplificationandoptionalPCR-freestepenabletheusertoachievereproducIBLyachievehighyieldsofDNAlibrarywithminimalsequencebiasandlowerrorrates.BackgroundInformationDNAlibrarypreparationisacriticalstepfornextgenerationsequencing(NGS).TogenerateaccuratesequencingdatainNGS,thepreparedlibraryDNAshouldbesufficientinyieldandofhighquality.AsNGStechnologyiscontinuouslyimproving,DNAlibrarypreparationisrequiredtobeoptimizedaccordingly. Mostofthecurrentlyusedmethodsaretime-consuming,expensive,inconvenient,andspecificallyneedlargeamountsofDNA.ThesefactorsresultinaDNAlibrarypreparationwhichcannotbeusedforBIOLOGicalsampleswithlimitedamountsofstartingmaterialsuchastumorbiopsy,earlyembryos,andembryonictissuesandcirculatingDNA.Inaddition,theamountofDNAenrichedbyChIPorMeDIP/hmeDIPisoftenatloworsub-nanogramlevelswhichcausesinsufficientDNAlibraryyields.Toaddressthisissue,EpigentekofferstheEpiNext™High-SensitivityDNALibraryPreparationKit.Principle&ProcedureThiskitincludesallreagentsrequiredateachstepoftheworkflowtocarryout asuccessfulDNAlibrarypreparation.Inthelibrarypreparation,DNAisfirstfragmentedtoanappropriatesize(about300bpsinpeaksize).Theendrepair/dAtailing(endpolishing)oftheDNAfragmentsareperformedsimultaneously.AdaptorsarethenligatedtobothendsofthepolishedDNAfragmentsforamplificationandsequencing.LigatedfragmentsaresizeselectedandpurifiedwithMQbeads,whichallowsforquickandprecisesizeselectionofDNA.Size-selectedDNAfragmentsareamplifiedwithahigh-fidelityPCRMixthatensuresmaximumyieldsfromminimumamountsofstartingmaterialandprovideshighlyaccurateamplificationoflibraryDNAwithlowerrorratesandminimumbias.StartingMaterialsStartingmaterialscanincludefragmenteddsDNAisolatedfromvarioustissueorcellsamples,dsDNAenrichedfromaChIPreaction,MeDIP/hMeDIPreaction,orexoncapture.DNAshouldberelativelyfreeofRNAbecauselargefractionsofRNAwillimpairendrepairanddA-tailing,resultinginreducedligationcapABIlities.TheinputamountofDNAcanbefrom0.2ngto100ng.Foroptimalpreparation,theinputamountshouldbe10ngto50ng.Fig.1. WorkflowoftheEpiNext™High-SensitivityDNALibraryPreparationKit.Fig.2. Sizedistributionoflibraryfragments.HumanplacentaDNAwasshearedtoaround300bpsinpeaksizeand0.2ngofDNAwasusedforDNAlibrarypreparationusingEpiNext™High-SensitivityDNALibraryPreparationKit.ProductComponents10XEndPolishingBuffer*EndPolishingEnzymeMix*EndPolishingEnhancer2XLigationBuffer*T4DNALigase*Adaptors(50µM)*MQBindingBeads*2XHiFiPCRMasterMix*PrimerU(10µM)*PrimerI(10µM)*ElutionBuffer*UserGuide*Spinthesolutiondowntothebottompriortouse.

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