f18ab大肠杆菌hcp基因缺失株构建及其对重要毒力因子分泌功能影响

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1、扬少I,1人学硕士学位论文差异。综上所述，hcp基因不影响该菌株以上五个重要毒力因二jIMIIIMIIIIHIIIHIHIl磊：11Y2258117移及到它们的分泌过程，尤其是Stx2e蛋白的分泌，所以引起了这些生物孚特翻减可藏丢失。关键词：斌ed重组系统；Ⅵ型分泌系统；hcp基因突变；粘附；侵袭；Vero细胞；毒力因子；stx2e周虹：F18ab+大肠杆菌hcp基因缺失株的构建及其对重要毒力冈子分泌功能～6⋯,h。,-N—Ant。hConstructionofF18ab-fimbriaedE．colihcpgenemutantandi

2、tseffectiononthesecretionofvirulancefactersAbstract[Background]EnterotoxigenicEscherichiacoli(STEC)strainF18abfrequentlycauseseverewaterydiarrhoeaandareresponsibleforsignificanteconomiclossesinthepigindustryworldwide．VirulencefactorsofSTECstrainF18abincludeadhesinssuchas

3、typeIfimbriae，F18abfimbriae，flagellum，AIDA(adhesininvolvedindiffuseadherence)andShigaliketoxin2e，whicharesecretedtoitsoutermembraneorsurroundingenvirenmentbysecretionsystem．Thusfar，sevensecretionsystemshavebeenidentified，namedTypeItoxypeⅦ，whichcarryouttheprocessoftranslo

4、cation，TypeVIandVIIbeingthemostrecentdiscovery．[PurposelTostudytheroleofhcpgeneofTypeⅥsecretionsysteminthesecretionofvirulancefactors．andfindnewwaystopreventandcontroltheSTEC—causeddisease．[MethodsandProcedures]BasedontheoriginalsequencesofhcpoperongeneclustersintheGenBa

5、nk，theSTECF18abwasselectedastheprototypeandconstructedthehcpmutantbyRedrecombinationsystem．First，wegeneratedPCRproductsbyprimerandsequencedofhcpgenefromSTECF18ab．Second，wegeneratedPCRproductsusingprimerswiththehomologiesextensionofhcpgenetobedeletedandtemplateplasmidpKD3

6、carryingselectableantibioticchloramphenicolresistance(cat)genethatisflankedbyFRT(FLPrecognitiontarget)sites．AndthenthePCRproductswereintroducedintoSTECF18abbyelectroporation，withthehelpofXRed—mediatedrecombinationsystem，hcpgenewasreplacedbythecatgene，thestrainsexpressing

7、chloramphenicolresistancegenewereselected，thechloramphenicolresistancegeneWaStheneliminatedbyusingahelperplasmid，pCP20，encodingtheFlprecombinase．Usingthissystem，hcpgeneinchromosomeofEscherichiacoliwasdeleted．STECF18abAhcpmutantobtainedwereconfirmedbyPCRamplificationandse

8、quencing．AndthenwecomparedtheSTECF18abandAhcpmutant、Ⅳiththeirbiologicalcharacteristics，includinggrowthr