receptor-mediated in vitro gene transformation by a soluble dna carrier system

receptor-mediated in vitro gene transformation by a soluble dna carrier system

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时间:2019-08-12

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1、THEJOURNALOFBIOLOGICALCHEHrSTRYVol.262No.10fasueofApril5pp.4429-44321987Q1987byThekmericanSocietyofBiciogicalChemis&.Ine.PrintedinU.S.A.Receptor-mediatedinVitroGenenalizedwithinhostcellsbyphagocytosis(2).Followinginter-nalization,afractionoftheDNAavoidsdigestionandeven-TransformationbyaSoluble

2、DNAtuallyentersthenucleusresultinginexpressionofnewgenesCarrierSystem*(1).Althoughtheprecipitationtechniqueiswidelyapplicableinvitro,itspotentialfordeliveryofgenesinvivoislimitedby(Receivedforpublication,December22,1986)alackofcellspecificityandespeciallybythe~~oL~~natureGeorgeY.Wu$andCatherin

3、eH.Wuoftheprecipitates.Wewonderedwhetherasimple,soluble,FromtheDivisionofGastroenterobgy-LiverDisease,DNAcarriersystemcouldbedevelopedtotargetDNAspe-DepartmentofMedicine,UniversityofConnecticutSchoolcificallytohepatocytesusingreceptor-mediatedendocytosis.ofMedicine,Farmington,Connecticut06032W

4、ebasedourexperimentsonthefollowingconcepts.I)Hepatocytespossessuniquereceptorsthatbindandinternal-Wepresent,here,evidencethatforeignDNAcanbeizegalactose-terminal(asialo-)glycoproteins(3).Couplingofspecificallydeliveredtocellsbyasolublecarriersys-DNAtoanasialoglycoproteincouldpermitinternalizat

5、ionoftemthattakesadvantageofreceptor-mediatedendo-DNAviaasialoglycoproteinreceptors.However,covalentCOU-cytosis.OurexperimentswerebasedonthefollowingplingmightaltertheDNAandprecludepropergeneexpres-concepts:1)hepatocytespossessauniquereceptorthatsion.2)Alternatively,DNAcanbeboundbypolycations,

6、e.g.bindsandinternalizesgalac~-terminal(asialo-)gIy-poly-L-lysineinastrong,electrostaticinteractionformingcoproteins;2)DNAcanbindtopolycationsinastrongsolublecomplexes(4).3)Chloramphenicolacetyltransferase,butnoncovaIentmannerformingsolublecomplexes;abacterialenzyme,catalyzesacetylationofthean

7、tibiotic,and3)thegeneforchloramphenicolacetyltransferase,chloramphenicol(5).Thegeneforthisenzymeisnotpresentabacterialenzymethatacetylateschloramphenicol,isinmammaliancells.notpresentinmammaliancells.Ourhypothesiswas:Ifpoly-L-lysinewere

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