Description: T4RNALigase2,alsoknownasT4Rnl2(gp24.1),catalyzesbothinter-andintramolecularformationofphosphodiesterbondsbetween5'-phosphateand3'-hydroxylterminiofoligonucleotides,single-strandedRNAandDNA.UnlikeT4RNALigase1,T4RNALigase2ismuchmoreactiveonjoiningnicksondoublestrandedRNAthanonjoiningtheendsofsingle-strandedRNA.Theenzymerequiresanadjacent5'-phosphateand3'-OHforligation.Theenzymecanalsoligatethe3'-OHofRNAtothe5'-phosphateofDNAinadoublestrandedstructure. Source: AnE.colistrainthatcarriestheT4RNALigase2gene(I.Schildkraut).
Application: -LigatesanickindsRNA -Ligatesthe3'-OHofRNAtothe5'-phosphateofDNAinadoublestrandedstructure
ReagentsSupplied: T4Rnl2ReactionBuffer
ReactionConditions: 1xT4Rnl2ReactionBuffer Incubateat37°C
10xT4RNALigase2ReactionBuffer: 500mMTris-HCl 20mMMgCl2 10mMDTT 4mMATP pH7.5@25°C
UnitDefinition: Oneunitisdefinedastheamountofenzymerequiredtoligate0.4µgofanequimolarmixofa23-merand17-merRNAsinatotalreactionvolumeof20µlin30minutesat37°C.
UnitAssayConditions: 1xT4Rnl2ReactionBufferand0.4µgofanequimolarmixofthe23-merand17-merRNAs.Afterincubationat37°Cfor30minutes,theligatedproductisdetectedona15%polyacrylamidegel. SpecificActivity:>40,000U/mg StorageConditions: 10mMTris-HCl 50mMKCl 35mM(NH4)2SO4 0.1mMDTT 0.1mMEDTA 50%Glycerol pH7.5@25°C
RecommendedStorageCondition:-20°C |