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Excellgen/ExcellScript Thermostable Reverse Transcriptase/EG-1029/40,000 Units

价格
¥1960.00
货号:EG-1029-40,000Units
浏览量:127
品牌:Excellgen
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商品描述
Description

ExCellscriptThermostableM-MuLVReverseTranscriptaseisaDNApolymerasewhichutilizesRNAasasubstrateandexhibitsnomeasurableproofreADIng3′→5′exonucleasefunction.ThisthermostablereversetranscriptasecanperformCDNAsynthesisreactionsthroughoutawiderangeoftemperaturesfrom42°Cto72°C.

Applications
  • FirststrandcDNAsynthesisforRT-PCRandreal-timeRT-PCR
  • SynthesisofcDNAforcloningandexpression
  • GenerationoflabeledcDNAprobesformicroarrays
  • DNAlabeling
  • AnalysisofRNAbyprimerextension
  • IsothermalnucleicacidamplificationsuchasLAMP
SourceArecombinantE.colistraincarryinganengineeredMoloney-MurineLeukemiaVirusReverseTranscriptasegene
QualityControl
  • Theabsenceofendo-,exodeoxyribonucleases,phosphatases,andribonucleasesconfirmedbyappropriatequalitytests.
  • FunctionallytestedinfirststrandcDNAsynthesis.
Unitdefinition

1unitisdefinedastheamountofenzymerequiredtoincorporate1nmolofdTTPintoacidinsolublematerialin10minutesat37°Cusingpolyr(A)/oligo(dT)asasubstrate.

Components
  • ThermostableM-MuLVReverseTranscriptase:200units/µLin50mMTris-HCl,150mMNaCl,0.1mMEDTA,1mMDTT,0.1%NP-40,50%glycerol,pH7.6@25°C
  • 10XReactionBuffer:500mMTris-HCl,750mMKCl,30mMMgCl2,100mMDTTpH8.3@25°C
Protocol
  1. PrimerAnnealing:CombinethefollowinginanRNase-freereactionvessel:
AmountDescriptionFinalConcentration
1µL25mMdNTPSolution(N2050L2.0mM
XµL1ng-2µgTotalRNA-or- 
XµL5-500ngmRNA(polyAselected) 
1µLOligo(dT)12-18(500µg/ml)-or-40µg/mL
1µLRandomPrimers(125µg/ml)-or10µg/mL
1µLGSPPrimer(2pmol)165µM
XµLSterileWaterN/A
10µLTotalVolume
  1. Heatreactionfor5minutesat65°C.Spinbriefly(5sec)topulldowncondensateandplaceimmediatelyonice.
  2. Add1µL10XM-MuLVRTBufferandDNase-freewatertoafinalvolumeof10µLperreaction.
  3. Incubate:
    • IfusingOligo(dT)orGSPprimers:2minutes@42°C
    • IfusingRandomprimers:2minutes@25°C
  4. Add1µL(200units)thermostableM-MuLVReverseTranscriptaseandmixbygentlypipettingsample.(Note:ifusingrandomprimers,pre-incubatereaction@25°Cfor10minutes).
  5. Incubateat42~72°Cfor45-60minutes.
  6. Inactivateenzymeat95°Cfor10minutes.
  7. Storeproductsat-20°Corproceedtonextstep.
Shipping4°C
NoteForresearchuseonly
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