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Oxford Genetics/pSF-RSV-Cas9WT (OG3537) CRISPR/Cas9 Expression Plasmid/OG3537/1 Ea
PlasmidInfo:
PlasmidInformation
ProductName:pSF-RSV-Cas9WT
ProductCode:OG3537
Size(bp):8227bp
BacterialAntibioticSelection:KanR
OriginandCompatibility:pUChighcopyderivedfrompBR322
BacterialCopyNumber:500-700percell
Promoter:RousSarcomaVirusLTRPromoter
PlasmidPurpose:
ThisplasmidisdesignedtoexpresswildtypeCas9inmammaliancells.ExpressionisdrivenfromaRSVpromoterinthemainMCS.
PromoterExpressionLevel:TheRousSarcomaLTR(RSV)promoterisbroadlyactiveinmanycelltypesandtissueinvivo,however,itisnotasstrongastheCMVpromoterinmostcelltypes.
SequenceandMap:
OtherInfo:
TranscriptionTermination:Thisplasmidcontainsthreealternativetranscriptionterminatorsformammalianbacterialandbacteriophage(T7)expression.Thismeansthatonlythepromoterneedstobechangedtoaltertheexpressionsystemyouareusing.Wesellmultiplepromotersthatcanbeusedineachofthesesystems.Thepresenceofeachterminatordoesnotreduceexpressioninthealternativesystems.
Cloning:
Cloninginagene:Thisplasmidcontainsagenewithinthemainmultiplecloningsite(NotI-ClaI).Anyplasmidthatwesellwherethegeneisthisconfigurationwillbelocatedintheexactsamepositioninrelationtothestartandstopcodonofthegeneintheplasmid.TheonlyexceptionstothisrulearefusionsproteinswherethefusiongenemaybepositionedatthefrontorendoftheMCStoallowgenefusion.
Bypositioningallofourgenesinthesamelocationitallowsthemtobetransferredbetweenplasmidsusingthesamecloningmethodandrestrictionsitesregardlessoftheplasmidbeingusedfromourproductrange.InsertinganewgeneintothisplasmidshouldbeeasilypossIBLeusingarangeofstandardrestrictionenzymesitesthatflankthegenecurrentlyinthevector.
Multiplecloningsitenotes:InthemultiplecloningsitetherearetwoimportantrestrictionsitescalledBsgIandBseRIsites.ThesesitesbothcuttheDNAatthesamepositionandcleavethestopcodonofthegeneinthemultiplecloningsiteinthisplasmidtherebyproducingaTAoverhang.Thisoverhangiscompatiblewithanyofourpeptideorreporterfusiontagplasmidsalsocutwitheitheroftheseenzymes.ThisallowsseamlessC-terminalfusionstobemadewiththegeneinthismultiplecloningsiteusingasinglecloningstepfromourC-terminalpeptideandreportertagproductrange.NormallytheeasiestmethodistoclonetheC-terminaltagfromourotherplasmidproductsintothisplasmidusingBsgI or BseRIandthedownstreamClaIrestrictionsite.
BseRIandBsgIsitesarenon-palindromicandcleaveadefinednumberofbasesawayfromtheirbindingsites.Thisallowsthemtocuttheupstreamstopcodoninthegeneinthisplasmidregardlessofthegenesequence.
IPStatus:
IntellectualPropertyStatusThisproductispartofourSnapFastProplasmidrange,formoreinformationontheIntellectualpropertystatusofthisplasmidpleaseclickhere
