PlasmidInfo:

PlasmidPurpose:

ThisvectoraddsaV5tagthatallowsproteindetectionorpurificationusingantibodiesagainsttheV5peptide.TheV5tagwillbepositionedatthe3primeend(Cterminus)ofagene(expressedprotein)thatiswithintheprimaryMCS(NotItoXbaI).TheV5tagcodingsequenceisGKPIPNPLLGLDST.Thereisanenterokinasecleavagesiteimmediatelyupstreamofthistag(DDDDK)thatcanbeusedtoremovethetagfromapurifiedproteinifrequired.Itcleavesafterthelysineresidue.

ThisplasmidcontainsthemammalianCMVpromotertodrivegeneexpression.WehavetestedallofourmammalianpromotersinarangeofcelltypesandCMVisconsistentlythestrongestinthosewehavestudied.HowevertherearemanyreportsoftheCMVpromoterdemonstratingsilencingbymethylationinlong-termculture.ForthisreasonwestockarangeofotherpromotersthatarecompatIBLewiththisplasmidandareavailableonrequest.

SequenceandMap:

OtherInfo:

Thisplasmidcontainsthreealternativetranscriptionterminatorsformammalianbacterialandbacteriophage(T7)expression.Thismeansthatonlythepromoterneedstobechangedtoaltertheexpressionsystemyouareusing.Wesellmultiplepromotersthatcanbeusedineachofthesesystems.Thepresenceofeachterminatordoesnotreduceexpressioninthealternativesystems.

Cloning:

ThereisastartcodonintheNcoIsitecanberemovedbydigestionwithKpnIifrequired.TheMCSforgeneinsertionsextendsfromNotItoXbaI.TherearebuiltinShine-DalgarnoandKOZAKsequencesalignedwiththestartcodonwithintheNcoIrestrictionsite.Iftheseareremovedyouwillneedtoaddaribosomalentrysitetothe5primeendofyourgeneforoptimalexpression.

IPStatus:

ThisproductispartofourSnapFastplasmidrange,formoreinformationontheIntellectualpropertystatusofthisplasmidpleaseclickhere

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