Details
Source
ChlorellavirusIL-3A
Note: Purifiedfromarecombinantsource(PatentNo.US005472872A)
ReagentsSupplied
10XCviJI*ReactionBuffer
DMSO
Description
- CviJI*isanuniquerestrictionenzymecapableofdigestingDNAattwoorthreebaserecognitionsequence(1,2)
- CviJI(Cat.No.2125-01)normallycleavesthesequence5"...PuGCPy...3"betweentheGandCtoleavebluntends
- Under"relaxed"conditions(inthepresenceofMg2+,ATPandenhancers),CviJI*cleavesthesequences5"...GC...3"except5"...PyGCPu...3"
- Capableofcleavingsingle-strandedDNAanddouble-strandedDNAintosmall20-200bpfragments
- GeneratesnumeroussequencespecificoligonucleotidesfromunknownDNAsamples
- ExclusivelyfromChimerx
Applications
- CviJIandCviJI*cleaveDNAextremelyfrequentlyandthuscanbeusedforavarietyofnovelmolecularBIOLOGyapplications(2,3,4)
- CviJI/CviJI*partialdigestscanalsobeusedinapplicationssuchasshot-guncloning,generatingquasi-randomlibraries(2)andepitopemappingorpanning
- CviJIdigestionofanonymousDNAproducesalargenumberofoligonucleotidesizedpolymersuponthermaldenaturation,whichcanbeexploitedinapplicationssuchas:
- Large-scalemappingorsequencingprojectsutilizinganonymousprimers
- HighresolutionmapppingofshortDNAs
- Nucleicacidlabeling(ThermalCycleLabeling,4,Fig.1)
- Detection(5)
- Amplification(5)
- Cloning(2,3)
- Captureofnucleicacids
IncubationTemperature
37°C
HeatInactivation
65°Cfor10minutes
AssayUnitSubstrate
pBR322
ReactionBuffer
10XCviJI*ReactionBuffer:
20mMglycyl-glycineKOH(pH8.5)
10mMmagnesiumacetate
50mMpotassiumacetate
0.1mMATP
0.1mMdithiothreitol
Note:100% DMSOsuppliedseparately
StorageBuffer
20mMTris-acetate(pH8.0at4°C)
0.5mMEDTA
0.1mMdithiothreitol
5mMmagnesiumchloride
50mMpotassiumacetate
50%(v/v)glycerol
AssayConditions
20mMglycylglycine-KOH(pH8.5)
10mMmagnesiumacetate
0.1mMdithiothreitol
50mMpotassiumacetate
0.1mMATP
30%DMSO
1µgpBR322
Incubationfor3hoursat37°Cinatotalreactionvolumeof25µl
StorageConditions
Storeat-80°C
Shippedondryice
CustomerNote(s)
(1)CviJI*restrictionendonucleaseisinhibitedbyglycerolconcentrationsinexcessof2.5%.ThereforetheextensionofthedigestiontimeisrecommendedratherthanusingadditionalunitsofCviJI*.Alternatively,DNAsamplecanbeethanolprecipitatedandre-digested.
(2)DuetoextremefrequencyofCviJI*/CviJIrecognitionsites,stericalinterferenceofcloselylocatedrecognitionsitesisobserved.Itresultsinslowerdigestionofsuchsites.Inconsequence,thegeneratedoligonucleotidefragmentsarerarelyshorterthan15bpthatmakesthemidealforanonymousprimerapplications.
(3)CviJI*reactionbuffercontainsDMSO,whichdoesnotinterferewithfurtherenzymaticmanipulations(ligations,labeling,etc).Ifthesampleisintendedforelectrophoresis,ethanolprecipitatationofthereactionmixtureaftercompleteddigestionisstronglyrecommendedinordertoavoiddiffusedbandsonagaroseorpolyacrylamidegels.
Downloads
CertificateofAnalysisPDF -CurrentLot
MSDSPDF -CurrentLot
References
(1)Xia,Y.,Burbank,D.,Uher,L.,Rabussay,D.andVanEtten,J.NucleicAcidsRes.15,6075-6090
(2)Fitzgerald,M.C.,Skowron,P.,VanEtten,J.L.,Smith,L.M.andMead,D.A.(1992)NucleicAcidsRes.20,3753-3762
(3)Skowron,P.M,Swaminathan,N.,McMaster,K.,George,D.,VanEtten,J.andMead,D.Gene157(1995)37-41
(4)Mead,D.,Swaminathan,N.,VanEttenJ.andSkowron,P.M.:RecombinantCviJIrestrictionendonuclease.(1995)UnitesStatesPatentnoUS005472872A
(5)Swaminathan,N.,McMaster,K.,Skowron,P.andMead,D.A.AnalyticalBiochemistry255(1998)133-141
