SIV Genome-Wide Pyrosequencing Provides a Comprehensive and Unbiased View of Variation within and outside CD8 T Lymphocyte Epitopes 英文参考文献.docVIP

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SIV Genome-Wide Pyrosequencing Provides a Comprehensive and Unbiased View of Variation within and outside CD8 T Lymphocyte Epitopes 英文参考文献.doc

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SIV Genome-Wide Pyrosequencing Provides a Comprehensive and Unbiased View of Variation within and outside CD8 T Lymphocyte Epitopes 英文参考文献

SIVGenome-WidePyrosequencingProvides aComprehensiveandUnbiasedViewofVariationwithin andoutsideCD8TLymphocyteEpitopes AustinL.Hughes1,ErickaA.Becker2,MichaelLauck3,JulieA.Karl2,AndrewT.Braasch3, DavidH.O’Connor2,3,ShelbyL.O’Connor2,3 * 1Department ofBiologicalSciences,University ofSouthCarolina, Columbia,South Carolina,United States ofAmerica, 2Wisconsin National PrimateResearchCenter, UniversityofWisconsin,Madison,Wisconsin,UnitedStatesofAmerica,3DepartmentofPathologyandLaboratoryMedicine,UniversityofWisconsin,Madison,Wisconsin, UnitedStatesofAmerica Abstract DeepsequencingtechnologyisrevolutionizingourunderstandingofHIV/SIVevolution.ItisknownthatacuteSIVsequence variationwithinCD8Tlymphocyte(CD8-TL)epitopesissimilaramongMHC-identicalanimals,butwedonotknowwhether thispersistsintothechronicphase.WenowdeterminewhetherchronicviralvariationinMHC-identicalanimalsinfected withclonalSIVissimilarthroughouttheentirecodingsequencewhenusingasensitivedeepsequencingapproach.We pyrosequencedtheentirecodingsequenceoftheSIVgenomeisolatedfromauniquecohortoffourSIVmac239-infected, MHC-identicalMauritiancynomolgusmacaques(MCM)48weeksafterinfection;oneMCMinthecohortbecameanelite controller.Amongthethreenon-controllers,wefoundthatgenome-widesequencesweresimilarbetweenanimalsandwe detectedincreasedsequencecomplexitywithin64%ofCD8-TLepitopeswhencomparedtoSangersequencingmethods. When we compared sequences between the MHC-matched controller and the three non-controllers, we found the viral population in the controller was less diverse and accumulated different variants than the viral populations in the non- controllers. Importantly, we found that initial PCR amplification of viral cDNA did not significantly affect the sequences detected,suggestingthatdataobtainedbypyrosequencingPCR-amplifiedviralcDNAaccuratelyrepresentsthediversityof sequences replicating within an animal. This demonstrates that chronic sequence diversity across the entire SIV coding sequence is similar among MHC-ident