Ligatesingle-strandedadenylatedDNAorRNAoligostoRNAtargetsforavarietyofapplications.

  • Epicentre ProductLigatesingle-strandedadenylatedDNAorRNAoligostoRNAtargets
  • UsethisenzymetoprepareCDNAlibrariesforsmall-RNAtranscriptomeanalysissuchasRNA-Seq
  • PerformlinkerligationformiRNAcloningapplications

T4RNALigase2,DeletionMutant,alsoknownasT4Rnl2(1-249),isusedtoligatesingle-strandedadenylatedDNAorRNAoligonucleotidestosmallRNAsforcloningornext-generationRNAsequencing.Thepreadenylated5´endsofDNAorRNAareligatedtothe3´endsofRNA.Unlikethefull-lengthenzyme,T4Rnl2(1-249)isunabletoadenylatethe5´endofthesubstrateinthepresenceofATP.However,itcanuseapreactivateddonor(App-DNAorApp-RNA)andjoinittothe3´endofanacceptor;thus,performingtheligationreactionintheabsenceofATPwhichpreventscircularizationandotherundesirablebimolecularreactions.

Theenzymeisavailablein2,000and10,000-Unitsizesataconcentrationof200U/µl.Theenzymeissuppliedwitha10XReactionBuffer.

UnitDefinition:Oneunitistheamountofenzymerequiredtogive50%ligationofa22-merRNAtothepreadenylatedendofa17-merDNAwhenbotholigosareannealedtoacomplementary39-merDNAstrandin30minutesat37°Cunderstandardassayconditions.

StorageBuffer:50%glycerolcontaining50mMTris-HCl(pH7.5),0.1MNaCl,0.1mMEDTA,1mMdithiothreitol(DTT),and0.1%Triton®X-100.

T4RNALigase2,DeletionMutant,10XReactionBuffer:500mMTris-HCl(pH7.5),20mMMgCl2,and10mMDTT.

ActivityAssay:Theunitdefinitionassayisperformedinareactioncontaining50mMTris-HCl(pH7.5),2mMMgCl2,1mMDTTand0.4µgofequimolarmixof22-mer,17-merand39-meroligonucleotides,andvaryingamountsofenzyme.

QualityControl:T4RNALigase2,DeletionMutant,isfreeofdetectableDNAexo-andendonuclease,andRNaseactivities.

ORDERINFORMATION

T4RNALigase2,DeletionMutantisprovidedat200U/μl.

Contents:T4RNALigase2DeletionMutant,10XReactionBuffer.