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NEB/HiScribe? T7 High Yield RNA Synthesis Kit/E2040S/50 reactions
  • NEB/HiScribe? T7 High Yield RNA Synthesis Kit/E2040S/50 reactions

NEB/HiScribe? T7 High Yield RNA Synthesis Kit/E2040S/50 reactions

價格: ¥2604.00 市場價: 4340.00

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    • Description:

      TheHiScribeT7HighYieldRNASynthesisKitisanextremelyflexIBLesystemforinvitrotranscriptionofRNAusingT7RNAPolymerase.ThekitallowsforsynthesismanykindsofRNAincludinginternallylabeledandco-transcriptionallycappedtranscripts.

      RNAsynthesizedfromthekitissuitableformanyapplicationsincludingRNAstructureandfunctionstudies,ribozymebiochemistry,probesforRNaseprotectionassaysandhybridizationbasedblots,anti-senseRNAandRNAiexperiments,microarrayanalysis,microinjection,andinvitrotranslationandRNAvaccines.

      Thekitcontainssufficientreagentsfor50reactionsof20μleach.Eachstandardreactionyieldsupto180μgofRNAfrom1μgcontroltemplate.Eachkitcanyieldupto9mgRNA.For32Plabeling,thekitcontainsenoughreagentsfor100?reactionsof20μleach.?

      MaterialsNotIncluded:
      • DNATemplate:TheDNAtemplatemustbelinearandcontaintheT7?RNAPolymerasepromoterwithcorrectorientationinrelationtotargetsequencetobetranscribed.?
      • CapAnalogs:NEB#S1411,#S1404,#S1405,#S1406and#S1407?
      • Modified-NTP:Biotin-,Fluorescein-,Digoxigenin-,orAminoallyl-NTP?
      • Labeling:[α-32P]labeledribonucleotide(800-6,000Ci/mmol)?
      • General:37°CincubatororPCRmachine,nuclease-freewater?
      • DNaseI:DNaseI(RNase-free)(NEB#M0303)?
      • Purification:Buffer-orwater-saturatedphenol/chloroform,ethanoland3Msodiumacetate,pH5.2,spincolumns?
      • GelAnalysis:Gelsandrunningbuffers,gelapparatus,powersupply

      Figure1.TranscriptionbyT7RNAPolymeraseFigure 1. Transcription by T7 RNA Polymerase

      Figure2.TimecourseofstandardRNAsynthesisfromthreeDNAtemplatesFigure 2. Time course of standard RNA synthesis from three DNA templates

      Reactionswereincubatedat37°CinaPCRmachine.TranscriptswerepurifiedbyspincolumnsandquantifiedonNanodrop?Spectrophotometer.
      Figure3.EffectoftemplateamountonRNAyieldFigure 3. Effect of template amount on RNA yield

      Standardreactionswereincubatedat37°CinaPCRmachinefor2hours.TranscriptswerepurifiedbyspincolumnsandquantifiedonNanoDrop?Spectrophotometer.
      Figure4:ImprovedRNAyieldandintegrityfromextendeddurationtranscriptionreactionsFigure 4: Improved RNA yield and integrity from extended duration transcription reactions

      reactionswereassembled,induplicate,accordingtothemanufacturers’suggestedprotocolsusing3ngofdsDNAtemplateencodinga1.8kbRNA,andincubatedat37°Cfor16,24and40hours.Ateachtimepoint,thecorrespondingtubesweretransferredto-20°Ctostopthereaction.Transcriptionreactionswerecolumnpurifiedafterthelasttimepoint.

      (A)Transcriptyield–Aftercolumnpurification,RNAconcentrationwasmeasuredusingaNanoDropspectrophotometerandtotalRNAyieldwascalculated.ThesedatademonstratethatasubstantiallyhigheryieldofRNAwassynthesizedusingtheHiScribeT7HighYieldRNASynthesisKitascomparedtothecompetitor’skit.

      (B)Transcriptintegrity–150ngofcolumnpurifiedRNAwasruna1.2%denaturingagarosegel,stainedwithethidiumbromideandvisualizedbyUVfluorescence.ThedatademonstrategreatlyimprovedtranscriptintegrityafterextendeddurationRNAsynthesisreactionsusingtheHiScribeT7HighYieldRNASynthesisKitascomparedtothecompetitor’skit.

      KitComponents

      Thefollowingreagentsaresuppliedwiththisproduct:

      Storeat(°C)Concentration
      ATP?-20100mM
      GTP?-20100mM
      UTP?-20100mM
      CTP?-20100mM
      10XT7ReactionBuffer-2010X
      FLucControlTemplate-200.5μg/μl
      T7RNAPolymeraseMix-20
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