Applications

• RemovalofgenomicDNAfromsmall-sampletotalRNApreparationsforexpressionanalysis(Fig.1)
• RemovalofsmallDNAoligonucleotides(e.g.,randomprimers)

Baseline-ZERO™DNase*digestsdsDNAandssDNAintomononucleotidesmoreeffectivelythanthecommonlyusedbovinepancreaticDNaseI.EventhesmallDNAoligonucleotidesthatremainaftertreatmentwithbovinepancreaticDNaseIareundetectablebygelelectrophoresisfollowingtreatmentwithBaseline-ZERODNase(Fig.2).RemovalofDNAfromRNApreparationsisparticularlybeneficialwhenRNAinasampleisamplifiedusingamethodthatinvolvesreversetranscriptionusingrandomprimers,sinceanycontaminatingDNAwouldalsobeatemplateforrandom-primedCDNAsynthesis.

Figure1.Real-timePCRofHeLaRNApreparationstreatedwithvariousDNases.ThelowertheC_Tvalue(intersectionofcurveswiththeredline),thegreatertheamountofresidualDNAnotdigestedbytheindicatedDNase.Thus,Baseline-ZERO™DNaseremovedalldetectableDNAfromtheRNAsample.TheTaqMan®probeassayamplifieda268-bpfragmentofβ-actin.Sampleswereruninduplicate.

UnitDefinition:OneMolecularBIOLOGyUnit(MBU)ofBaseline-ZERO™DNaseproducesanincreaseintheA₂₆₀ofasolutionofdsDNA,of0.001perminuteat25°C.Functionally,1MBUcompletelydigests1µgoflinearpUC19DNAtomononucleotidesin10minutesat37°C.

StorageBuffer:Baseline-ZERODNaseissuppliedina50%glycerolsolutioncontaining50mMTris-HCl(pH7.5),10mMCaCl₂,10mMMgCl₂and0.1%Triton®X-100.

10XBaseline-ZERO™DNaseReactionBuffer:100mMTrisHCl(pH7.5),25mMMgCl₂and5mMCaCl₂.

10XBaseline-ZERO™DNaseStopSolution:30mMEDTA.

QualityControl:Baseline-ZERODNaseisassayedforitsABIlitytocompletelydigestlineardsDNAtomononucleotidesunderstandardassayconditions.Baseline-ZERODNaseisfreeofdetectableRNaseactivitiesasassayedbyPAGEanalysisof1µgofasyntheticRNAtranscriptfollowinganovernightincubationwithsufficientDNaseItocompletelydigest1,000 µgofDNA.

References

1. Kienzle,N.etal.(1996)BioTechniques20,612.

*Patentpending.