Details
Description
- Thermostableenzymeofapproximately94kDafrom Thermusaquaticus
- Ultrapure,Recombinantprotein
Applications
- RecommendedforuseinPCRandprimerextensionreactionsatelevatedtemperaturestoobtainawiderangeofDNAproducts upto10Kb
- ReplicatesDNAat74°Candexhibitsahalf-lifeof40minutesat95°C(1,2)
- CatalyzesthepolymerizationofnucleotidesintoduplexDNAinthe5"→3"directioninthepresenceofmagnesiumions
- Maintainsthe5"→3"exonucleaseactivity
- Lacksthe3"→5"exonucleaseactivity
- AddsextraAatthe3"-ends
Advantages
- Helpsvisualizetheadditionofthepolymerasetothereaction
- Confirmscompletemixing
- EnablesdirectloADIngofPCRproductsontoanagarosegelwithouttheadditionofagelloadingbuffer
- Addeddyesallowtrackingofelectrophoresisprogress
- DoesnotinterferewithPCRperformance
- Doesnotinterferewithmostdownstreamapplications
Note:Notrecommendedforanyapplicationsusingabsorbanceorfluorescenceexcitation
ReagentsSupplied
10XTaqBufferA
10XTaqBufferB
UnitDefinition
Oneunitisdefinedastheamountofenzymerequiredtocatalyzetheincorporationof10nmolesofdNTPintoacid-insolublematerialat74°Cin30minutes
ReactionBuffer
10xTaqBufferA(optimizationbufferwithoutMgCl2): allowstooptimizeMgCl2 concentration
10xTaqBufferB(generalapplication,upto10kb): contains15mM MgCl2 andis optimizedforusewith0.2 mMofeachdNTP
StorageBuffer
20mMTris-HCl(pH8.0at22°C
100mMKCl
0.1mMEDTA
1mMdithiothreitol
50%(v/v)glycerol
Stablizers
AssayConditions
50mMTris-HCl(pH9.0at25°C)
50mMNaCl
50mMMgCl2
200µMeachofdATP,dCTP,dGTPanddTTP(amixofunlabeledand[3H]dTTP
10µgactivatedcalfthymusDNA
0.1mg/mlbovineserumalbumin
Finalvolumeof50µl
QualityControl
Allpreparationsareassayedforcontaminatingendonuclease,3"-exonuclease,andnonspecificsingle-anddouble-strandedDNaseactivities
Typicalpreparationsaregreaterthan95%pure,asjudgedbySDSpolyacrylamidegelelectrophoresis
StorageConditions
Storeat-20°C
Productshippedondryice
References
1.Chien,A.,Edgar,D.B.andTrela,J.M.(1976)J.Bacteriol.127,1550
2.Kaledin,A.S.,Sliusarenko,A.G.andGorodetskii,S.I.(1980)Biokhimiya45,644
