简介：Expressedsequencetags(ESTs)arewidelyusedingenesurveyresearchtheseyears.TheESTPipelineSystem,softwaredevelopedbyHangzhouGenomicsInstitute(HGI),canautomaticallyanalyzedifferentscalarESTsequencesbysuitablemethods.Alltheanalysisreports,includingthoseofvectormasking,sequenceassembly,geneannotation,GeneOntologyclassification,andsomeotheranalyses,canbebrowsedandsearchedaswellasdownloadedintheExcelformatfromthewebinterface,savingresearcheffortsfromroutinedataprocessingforbiologicalrulesembeddedinthedata.

简介：AnnotationofthegenomesequenceoftheSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus)isindispensabletounderstanditsevolutionandpathogenesis.WehaveperformedafullannotationoftheSARS-CoVgenomesequencesbyusingannotationprogramspubliclyavailableordevelopedbyourselves.Totally,21openreadingframes(ORFs)ofgenesorputativeuncharacterizedproteins(PUPs)werepredicted.SevenPUPshadnotbeenreportedpreviously,andtwoofthemwerepredictedtocontaintransmembraneregions.EightORFspartiallyoverlappedwithorembeddedintothoseofknowngenes,revealingthattheSARS-CoVgenomeisasmallandcompactonewithoverlappedcodingregions.ThemoststrikingdiscoveryisthatanORFlocatesontheminusstrand.Wehavealsoannotatednon-codingregionsandidentifiedthetranscriptionregulatingsequences(TRS)intheintergenicregions.TheanalysisofTRSsupportstheminusstrandextendingtranscriptionmechanismofcoronavirus.TheSNPanalysisofdifferentisolatesrevealsthatmutationsofthesequencesdonotaffectthepredictionresultsofORFs.

简介：Knowledgeoftheevolutionofpathogensisofgreatmedicalandbiologicalsignificancetotheprevention,diagnosis,andtherapyofinfectiousdiseases.InordertounderstandtheoriginandevolutionoftheSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus),wecollectedcompletegenomesequencesofallvirusesavailableinGenBank,andmadecomparativeanalyseswiththeSARSCoV.GenomicsignatureanalysisdemonstratesthatthecoronavirusesalltaketheTGTTastheirrichesttetranucleotideexcepttheSARS-CoV.Adetailedanalysisoftheforty-twocompleteSARS-CoVgenomesequencesrevealedtheexistenceoftwodistinctgenotypes,andshowedthattheseisolatescouldbeclassifiedintofourgroups.OurmanualanalysisoftheBLASTNresultsdemonstratesthattheHE(hemagglutinin-esterase)geneexistsintheSARS-CoV,andmanymutationsmadeitunfamiliartous.

简介：Inthefaceoftheworldwidethreatofsevereacuterespiratorysyndrome(SARS)tohumanlife,someofthemosturgentchallengesaretodevelopfastandaccurateanalyticalmethodsforearlydiagnosisofthisdiseaseaswellastocreateasafeanti-viralvaccineforprevention.Totheseends,weinvestigatedtheantigenicityofthespikeprotein(Sprotein),amajorstructuralproteinintheSARS-coronavirus(SARS-CoV).BaseduponthetheoreticalanalysisforhydrophobicityoftheSprotein,18peptidesweresynthesized.UsingEnzyme-LinkedImmunosorbentAssay(ELISA),thesepeptideswerescreenedintheserafromSARSpatients.Accordingtotheseresults,twofragmentsoftheSgenewereamplifiedbyPCRandclonedintopET-32a.BothSfragmentswereexpressedintheBL-21strainandfurtherpurifiedwithanaffinitychromatography.TheserecombinantSfragmentswereconfirmedtohavepositivecross-reactionswithSARSsera,eitherbyWesternblotorbyELISA.OurresultsdemonstratedthatthepotentialepitoperegionswerelocatedatCodons469-882intheSprotein,andoneepitopesitewaslocatedatCodons599-620.IdentificationofantigenicregionsintheSARS-CoVSproteinmaybeimportantforthefunctionalstudiesofthisvirusorthedevelopmentofclinicaldiagnosis.

简介：Thenucleocapsidprotein(Nprotein)hasbeenfoundtobeanantigenicproteininanumberofcoronaviruses.WhethertheNproteininsevereacuterespiratorysyndrome-associatedcoronavirus(SARS-CoV)isantigenicremainstobeelucidated.UsingWesternblotandEnzyme-linkedImmunosorbentAssay(ELISA),therecombinantNproteinsandthesynthesizedpeptidesderivedfromtheNproteinwerescreenedinserafromSARSpatients.AllpatientserainthisstudydisplayedstrongpositiveimmunoreactivitiesagainsttherecombinantNproteins,whereasnormalseragavenegativeimmunoresponsestotheseproteins,indicatingthattheNproteinofSARS-CoVisanantigenicprotein.Furthermore,theepitopesitesintheNproteinweredeterminedbycompetitionexperiments,inwhichtherecombinantproteinsorthesynthesizedpeptidescompetedagainsttheSARS-CoVproteinstobindtotheantibodiesraisedinSARSsera.OneepitopesitelocatedattheC-terminuswasconfirmedasthemostantigenicregioninthisprotein.AdetailedscreeningofpeptidewithELISAdemonstratedthattheaminosequencefromCodons371to407wastheepitopesiteattheC-terminusoftheNprotein.UnderstandingoftheepitopesitescouldbeverysignificantfordevelopinganeffectivediagnosticapproachtoSARS.

简介：InordertodevelopclinicaldiagnostictoolsforrapiddetectionofSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus)andtoidentifycandidateproteinsforvaccinedevelopment,theC-terminalportionofthenucleocapsid(NC)genewasamplifiedusingRT-PCRfromtheSARS-CoVgenome,clonedintoayeastexpressionvector(pEGH),andexpressedasaglutathioneS-transferase(GST)andHisx6double-taggedfusionproteinunderthecontrolofaninduciblepromoter.WesternanalysisonthepurifiedproteinconfirmedtheexpressionandpurificationoftheNCfusionproteinsfromyeast.Todetermineitsantigenicity,thefusionproteinwaschallengedwithserumsamplesfromSARSpatientsandnormalcontrols.TheNCfusionproteindemonstratedhighantigenicitywithhighspecificity,andtherefore,itshouldhavegreatpotentialindesigningclinicaldiagnostictoolsandprovideusefulinformationforvaccinedevelopment.

简介：BeijinghasbeenoneoftheepicentersattackedmostseverelybytheSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus)sincethefirstpatientwasdiagnosedinoneofthecity'shospitals.WenowreportcompletegenomesequencesoftheBJGroup,includingfourisolates(IsolatesBJ01,BJ02,BJ03,andBJ04)oftheSARS-CoV.ItisremarkablethatallmembersoftheBJGroupshareacommonhaplotype,consistingofsevenlocithatdifferentiatethegroupfromotherisolatespublishedtodate.Among42substitutionsuniquelyidentifledfromtheBJgroup,32arenon-synonymouschangesattheaminoacidlevel.Rootedphylogenetictrees,proposedonthebasisofhaplotypesandothersequencevariationsofSARS-CoVisolatesfromCanada,USA,Singapore,andChina,gaverisetodifferentparadigmsbutpositionedtheBJGroup,togetherwiththenewlydiscoveredGD01(GD-Ins29)inthesameclade,followedbytheH-UGroup(fromHongKongtoUSA)andtheH-TGroup(fromHongKongtoToronto),leavingtheSPGroup(Singapore)moredistant.ThisresultappearstosuggestapossibletransmissionpathfromGuangdongtoBeijing/HongKong,thentoothercountriesandregions.

简介：Werecentlyreportedtheuseofagene-trappingapproachtoisolatecellclonesinwhichareportergenehadintegratedintogenesmodulatedbyT-cellactivation.WehavenowtestedapanelofclonesfromthatreportandidentifiedtheonethatrespondstoavarietyofG-proteincoupledreceptors(GPCR).TheβlactamasetaggedEGR-3JurkatcellwasusedtodissectspecificGPCRsignalinginvivo.ThreeGPCRswerestudied,includingthechemokinereceptorCXCR4(Gicoupled)thatwasendogenouslyexpressed,theplateletactivationfactor(PAF)receptor(Gq-coupled),andβ2adrenergicreceptor(Gs-coupled)thatwasbothstablytransfected.Agonistsforeachreceptoractivatedtranscriptionoftheβ-lactamasetaggedEGR-3gene.InductionofEGR-3throughCXCR4wasblockedbypertussistoxinandPD58059,aspecificinhibitorofMEK(MAPK/ERKkinase).NeitheroftheseinhibitorsblockedisoproterenolorPAF-mediatedactivationofEGR-3.Conversely,β2-andPAF-mediatedEGR-3activationwasblockedbythep38,specificinhibitorSB580.Inaddition,bothβ2-andPAF-mediatedEGR-3activationcouldbesynergisticallyactivatedbyCXCR4activation.ThiscombinedresultindicatesthatEGR-3canbeactivatedthroughdistinctsignaltransductionpathwaysbydifferentGPCRsandthatsignalscanbeintegratedandamplifiedtoefficientlytunethelevelofactivation.

简介：obtainaninitialoverviewofgenediversityandexpressionpatterninporcinethymus,11,712ESTs(ExpressedSequenceTags)from100-day-oldporcinethymus(FTY)weresequencedand7,071cleanedESTswereusedforgeneexpressionanalysis.ClusteredbythePHRAPprogram,959contigsand3,074singletswereobtained.Blastsearchshowedthat806contigsand1,669singlets(totally5,442ESTs)hadhomologuesinGenBankand1,629ESTswerenovel.AccordingtotheGeneOntologyclassification,36.99%ESTswerecatalogedintothegeneexpressiongroup,indicatingthatalthoughthefunctionalgene(18.78%indefensegroup)ofthymusisexpressedinacertaindegree,the100-day-oldporcinethymusstillexistsinadevelopmentalstage.Comparativeanalysisshowedthatthegeneexpressionpatternofthe100-day-oldporcinethymusissimilartothatofthehumaninfantthymus.

简介：Nineshorttandemrepeat(STR)markers(D3S1358,VWA,FGA,THO1,TPOX,CSFIPO,D5S818,D13S317,andD7S820)andasex-identificationmarker(Amel-ogeninlocus)wereamplifiedwithmultiplexPCRandweregenotypedwithafour-colorfluorescencemethodinsamplesfrom174unrelatedHanindividualsinNorthChina.Theallelefrequencies,genotypefrequencies,heterozygosity,prob-abilityofdiscriminationpowers,probabilityofpaternityexclusionandHardy-Weinbergequilibriumexpectationsweredetermined.TheresultsdemonstratedthatthegenotypesatalltheseSTRlociinHanpopulationconformtoHardy-Weinbergequilibriumexpectations.Thecombineddiscriminationpower(DP)was1.05×10-10withinnineSTRlocianalyzedandtheprobabilityofpaternityexclusion(EPP)was0.9998.TheresultsindicatethatthesenineSTRlociandtheAmelo-geninlocusareusefulmarkersforhumanidentification,paternityandmaternitytestingandsexdeterminationinforensicsciences.

简介：TheR(replicase)proteinistheuniquelydefinednon-structuralprotein(NSP)responsibleforRNAreplication,mutationrateorfidelity,regulationoftranscrip-tionincoronavirusesandmanyotherssRNAviruses.Basedonourcompletegenomesequencesoffourisolates(BJ01-BJ04)ofSARS-CoVfromBeijing,China,weanalyzedthestructureandpredictedfunctionsoftheRproteinincomparisonwith13otherisolatesofSARS-CoVand6othercoronaviruses.TheentireORF(open-readingframe)encodesfortwomajorenzymeactivities,RNA-dependentRNApolymerase(RdRp)andproteinaseactivities.TheRpolyproteinunder-goesacomplexproteolyticprocesstoproduce15function-relatedpeptides.Ahydrophobicdomain(HOD)andahydrophilicdomain(HID)arenewlyidentifiedwithinNSP1.ThesubstitutionrateoftheRproteinisclosetotheaverageoftheSARS-CoVgenome.ThefunctionaldomainsinallNSPsoftheRproteingivedifferentphylogeneticresultsthatsuggesttheirdifferentmutationrateunderselectivepressure.ElevenhighlyconservedregionsinRdRpandtwelvecleavagesitesby3CLP(chymotrypsin-likeprotein)havebeenidentifiedaspotentialdrugtargets.Findingssuggestthatitispossibletoobtaininformationaboutthephy-logenyofSARS-CoV,aswellaspotentialtoolsfordrugdesign,genotypinganddiagnosticsofSARS.

简介：TheCoronaviridaefamilyischaracterizedbyanucleocapsidthatiscomposedofthegenomeRNAmoleculeincombinationwiththenucleoprotein(Nprotein)withinavirion.ThemoststrikingphysiochemicalfeatureoftheNproteinofSARS-CoVisthatitisatypicalbasicproteinwithahighpredictedpIandhighhydrophilicity,whichisconsistentwithitsfunctionofbindingtotheribophosphatebackboneoftheRNAmolecule.ThepredictedhighextentofphosphorylationoftheNproteinonmultiplecandidatephosphorylationsitesdemonstratesthatitwouldberelatedtoimportantfunctions,suchasRNA-bindingandlocalizationtothenucleolusofhostcells.SubsequentstudyshowsthatthereisanSR-richregionintheNproteinandthisregionmightbeinvolvedintheprotein-proteininteraction.TheabundantantigenicsitespredictedintheNprotein,aswellasexperimentalevidencewithsynthesizedpolypeptides,indicatethattheNproteinisoneofthemajorantigensoftheSARS-CoV.Comparedwithotherviralstructuralproteins,thelowvariationrateoftheNproteinwithregardstoitssizesuggestsitsimportancetothesurvivalofthevirus.

简介：TheE(envelope)proteinisthesmalleststructuralproteininallcoronavirusesandistheonlyviralstructuralproteininwhichnovariationhasbeendetected.WeconductedgenomesequencingandphylogeneticanalysesofSARS-CoV.Basedongenomesequencing,wepredictedtheEproteinisatransmembrane(TM)pro-teincharacterizedbyaTMregionwithstronghydrophobicityandα-helixcon-formation.Weidentifiedasegment(NH2-_L-Cys-A-Y-Cys-Cys-N_-COOH)inthecarboxyl-terminalregionoftheEproteinthatappearstoformthreedisulfidebondswithanothersegmentofcorrespondingcysteinesinthecarboxyl-terminusoftheS(spike)protein.ThesebondspointtoapossiblestructuralassociationbetweentheEandSproteins.OurphylogeneticanalysesoftheEproteinsequencesinallpub-lishedcoronavirusesplaceSARS-CoVinanindependentgroupinCoronaviridaeandsuggestanon-humananimalorigin.

简介：WestudiedstructuralandimmunologicalpropertiesoftheSARS-CoVM(mem-brane)protein,basedoncomparativeanalysesofsequencefeatures,phylogeneticinvestigation,andexperimentalresults.TheMproteinispredictedtocontainatriple-spanningtransmembrane(TM)region,asingleN-glycosylationsitenearitsN-terminusthatisintheexteriorofthevirion,andalongC-terminalregionintheinterior.TheMproteinharborsahighersubstitutionrate(0.6%correlatedtoitssize)amongviralopenreadingframes(ORFs)frompublisheddata.ThefoursubstitutionsdetectedintheMprotein,whichcausenon-synonymouschanges,canbeclassifiedintothreetypes.OneofthemresultsinchangesofpI(isoelectricpoint)andcharge,affectingantigenicity.ThesecondchangeshydrophobicityoftheTMregion,andthethirdonerelatestohydrophilicityoftheinteriorstructure.PhylogenetictreebuildingbasedonthevariationsoftheMproteinappearstosupportthenon-humanoriginofSARS-CoV.Toinvestigateitsimmunogenicity,wesynthesizedeightoligopeptidescovering69.2%oftheentireORFandscreenedthembyusingELISA(enzyme-linkedimmunosorbentassay)withserafromSARSpatients.Theresultsconfirmedourpredictionsonantigenicsites.

简介：Thecorona-likespikesorpeplomersonthesurfaceofthevirionunderelectronicmicroscopearethemoststrikingfeaturesofcoronaviruses.TheS(spike)proteinisthelargeststructuralprotein,with1,255aminoacids,intheviralgenome.Itsstructurecanbedividedintothreeregions:alongN-terminalregionintheexte-rior,acharacteristictransmembrane(TM)region,andashortC-terminusintheinteriorofavirion.WedetectedfifteensubstitutionsofnucleotidesbycomparisonswiththeseventeenpublishedSARS-CoVgenomesequences,eight(53.3%)ofwhicharenon-synonymousmutationsleadingtoaminoacidalternationswithpredictedphysiochemicalchanges.ThepossibleantigenicdeterminantsoftheSproteinarepredicted,andtheresultisconfirmedbyELISA(enzyme-linkedimmunosorbentassay)withsynthesizedpeptides.AnotherprofoundfindingisthatthreedisulfidebondsaredefinedattheC-terminuswiththeN-terminusoftheE(envelope)pro-tein,basedonthetypicalsequenceandpositions,thusestablishingthestructuralconnectionwiththesetwoimportantstructuralproteins,ifconfirmed.Phyloge-neticanalysisrevealsseveralconservedregionsthatmightbepotentdrugtargets.

简介：Wereportacompletegenomicsequenceofrareisolates(minorgenotype)oftheSARS-CoVfromSARSpatientsinGuangdong,China,wherethefirstfewcasesemerged.Themoststrikingdiscoveryfromtheisolateisanextra29-nucleotidesequencelocatedatthenucleotidepositionsbetween27,863and27,864(referredtothecompletesequenceofBJ01)withinanoverlappedregioncomposedofBGI-PUP5(BGI-postulateduncharacterizedprotein5)andBGI-PUP6upstreamoftheN(nucleocapsid)protein.Thediscoveryofthisminorgenotype,GD-Ins29,suggestsasignificantgeneticeventanddifferentiatesitfromthepreviouslyre-portedgenotype,thedominantformamongallsequencedSARS-CoVisolates.A17-ntsegmentofthisextrasequenceisidenticaltoasegmentofthesamesizeintwohumanmRNAsequencesthatmayinterferewithviralgenomereplicationandtranscriptioninthecytosoloftheinfectedcells.Itprovidesanewavenuefortheexplorationofthevirus-hostinteractioninviralevolution,hostpathogenesis,andvaccinedevelopment.

简介：MITEs(Miniatureinverted-repeattransposableelements)arereminiscenceofnon-autonomousDNA(classⅡ)elements,whicharedistinguishedfromothertranspos-ableelementsbytheirsmallsize,shortterminalinvertedrepeats(TIRs),highcopynumbers,genicpreference,andDNAsequenceidentityamongfamilymembers.Al-thoughMITEswerefirstdiscoveredinplantsandstillactivelyreshapinggenomes,theyhavebeenisolatedfromawiderangeofeukaryoticorganisms.MITEscanbedividedintoTourist-like,Stowaway-like,andpogo-likegroups,accordingtosimilaritiesoftheirTIRsandTSDs(targetsiteduplications).Indespiteofsev-eralmodelstoexplaintheoriginandamplificationofMITEs,theirmechanismsoftranspositionandaccumulationineukaryoticgenomesremainpoorlyunderstoodowingtoinsufficientexperimentaldata.TheuniquepropertiesofMITEshavebeenexploitedasusefulgenetictoolsforplantgenomeanalysis.UtilizationofMITEsaseffectiveandinformativegenomicmarkersandpotentialapplicationofMITEsinplantssystematic,phylogenetic,andgeneticstudiesarediscussed.

简介：Theterms"science"(HanyuPinyin:Kēxué)and"technology"(Jìshù),inwesternlanguages,areusu-allyusedseparatelywithdifferentmeaningsorto-getheras"scienceandtechnology".However,theyareoftenmentionedas"KēxuéJìshù",orevenabbre-viatedas"sci-tech(Kējì)"inChina(ItisusedinasimilarwayinJapan).

简介：Thestrongestsignalofplantpromoterissearchedwiththemodelofsinglemotifwithtwotypes.ItturnsoutthatthedominanttypeistheTATA-box.TheothertypemaybecalledTATA-lesssignal,andmaybeusedingenefindersforpromoterrecognition.WhiletheTATAsignalsareverycloseforthemonocotandthedicot,theirTATA-lesssignalsaresignificantlydifferent.Ageneralandflexiblemulti-motifmodelisalsoproposedforpromoteranalysisbasedondynamicprogramming.ByextendingtheGibbssamplertothedynamicprogrammingandintroducingtemperature,anefficientalgorithmisdevelopedforsearchingsignalsinplantpromoters.

简介：Differentialproteomeprofilesofhumanlungsquamouscarcinomatissuecomparedtopairedtumor-adjacentnormalbronchialepithelialtissuewereestablishedandanalyzedbymeansofimmobilizedpHgradient-basedtwo-dimensionalpolyacrylamidegelelectrophoresis(2-DPAGE)andmatrix-assistedlaserdesorption/ionizationtimeofflightmassspectrometry(MALDI-TOF-MS).Theresultsshowedthatwell-resolved,reproducible2-DEpatternsofhumanlungsquamouscarcinomaandadjacentnormalbronchialepithelialtissueswereobtainedundertheconditionof0.75-ugprotein-load.Theaveragedeviationofspotpositionwas0.733+0.101mminIEFdirection,and0.925+0.207mminSDS-PAGEdirection.Fortumortissue,atotalof1241±88spotsweredetected,987±65spotswerematchedwithanaveragematchingrateof79.5%.Forcontrol,atotalof1190+72spotsweredetected,and875±48spotswerematchedwithanaveragematchingrateof73.5%.Atotalof864±34spotswerematchedbetweentumorsandcontrols.Forty-threedifferentialproteinswerecharacterized:someproteinswererelatedtooncogenes,andothersinvolvedintheregulationofcellcycleandsignaltransduction.Itissuggestedthatthedifferentialproteomicapproachisvaluableformassidentificationofdifferentiallyexpressedproteinsinvolvedinlungcarcinogenesis.Thesedatawillbeusedtoestablishhumanlungcancerproteomedatabasetofurtherstudyhumanlungsquamouscarcinoma.