Baculovirus Superinfection A Probable Restriction Factor on the Surface Display of Proteins for Library Screening.docVIP

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Baculovirus Superinfection A Probable Restriction Factor on the Surface Display of Proteins for Library Screening.doc

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Baculovirus Superinfection A Probable Restriction Factor on the Surface Display of Proteins for Library Screening

BaculovirusSuperinfection:AProbableRestrictionFactor ontheSurfaceDisplayofProteinsforLibraryScreening XiaodongXu1,YuanrongChen1,YuZhao1,XiaofenLiu1,BeitaoDong1,IanM.Jones2,HongyingChen1* 1CollegeofLifeSciences,NorthwestAFUniversity,Yangling,Shaanxi,P.R.China,2SchoolofBiologicalSciences,UniversityofReading,Reading,UnitedKingdom Abstract Inadditiontotheexpressionofrecombinantproteins,baculoviruseshavebeendevelopedasaplatformforthedisplayof complex eukaryotic proteins on the surface of virus particles or infected insect cells. Surface display has been used extensivelyforantigenpresentationandtargetedgenedeliverybutisalsoacandidateforthedisplayofproteinlibrariesfor molecularscreening.However,althoughbaculovirusgenelibrariescanbeefficientlyexpressedanddisplayedonthesurface ofinsectcells,targetgeneselectionisinefficientprobablyduetosuper-infectionwhichgivesrisetocellsexpressingmore thanoneprotein.InthisreportbaculovirussuperinfectionofSf9cellshasbeeninvestigatedbytheuseoftworecombinant multiplenucleopolyhedroviruscarryinggreenorredfluorescentproteinsunderthecontrolofbothearlyandlatepromoters (vAcBacGFPandvAcBacDsRed).Thereportergeneexpressionwasdetected8hoursaftertheinfectionofvAcBacGFPand cells in early and late phases of infection could be distinguished by the fluorescence intensity of the expressed protein. Simultaneous infection with vAcBacGFP and vAcBacDsRed viruses each at 0.5 MOI resulted in 80% of infected cells co- expressing the two fluorescent proteins at 48 hours post infection (hpi), and subsequent infection with the two viruses resultedinsimilarco-infectionrate.MostSf9cellswerere-infectablewithinthefirstseveralhourspostinfection,butthere- infectionratethendecreasedtoaverylowlevelby16hpi.OurdatademonstratethatSf9cellswereeasilysuper-infectable during baculovirus infection, and super-infection could occur simultaneously at the time of the primary infection or subsequentlyduringsecondaryinfectionbyprogenyviruses.Theefficiencyofsuper-infectionmayexplainthediff