Objective:TostudythebiologicalactivityofMycoplasmapenetrans35kDalipoprotein(P35)invitro,prokaryoticexpressionvectorpQE31/p35wasconstructedandrecombinantfusionproteinP35(rP35)wasexpressedinE.coli.Methods:Thep35genewasamplifiedbypolymerasechainreaction(PCR),clonedtopQE31,andapositiveclonewasscreened.PCR-mediatedmutagenesiswasusedtochangethetwo"TGA"tripletsto"TGG"tripletswithinthep35gene.ProductionoftherecombinantproteinwasinducedbytheadditionofIPTGtotheE.coliculture,rP35waspurifiedwithaNi-NTASpinKitandrP35purificationwasanalyzedbyWesternblot.Results:About1KbPCRamplificationwasclonedintopQE31.Thetwo"TGA"tripletswithinthep35geneweresuccessfullychangedto"TGG"triplets.ThepQE31/p35vectorexpressedaproteinwithacalculatedmolecularmassof37.4kDainE.coli.Westernblotindicatedthe37.4kDaproteinwasrP35.Conclusion:PQE311p35,aprokaryoticexpressionvectorcontainingp35gene,wassuccessfullyconstructedandexpressedinE.coli.
