High-resolution human genome structure by single-molecule ...

This report presents the analysis of structural variation in four human genomes using Optical Mapping, compares these results to other genome- ... Skiptomaincontent PNASJune15,2010107(24)10848-10853;https://doi.org/10.1073/pnas.0914638107 BrianTeagueFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite MichaelS.WatermanFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite StevenGoldsteinFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite KonstantinosPotamousisFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite ShiguoZhouFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite SusanReslewicFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite DeepayanSarkarFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite AntonValouevFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite ChristopherChurasFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite JeffreyM.KiddFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite ScottKohnFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite RodneyRunnheimFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite CaseyLamersFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite DanForrestFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite MichaelA.NewtonFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite EvanE.EichlerFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite MarijoKent-FirstFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite UrvashiSurtiFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite MironLivnyFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite DavidC.SchwartzFindthisauthoronGoogleScholar FindthisauthoronPubMed Searchforthisauthoronthissite Forcorrespondence: [email protected] Edited*byDavidE.Housman,MassachusettsInstituteofTechnology,Cambridge,MA,andapprovedMay6,2010(receivedforreviewDecember17,2009) Article Figures&SI Info&Metrics PDF AbstractVariationingenomestructureisanimportantsourceofhumangeneticpolymorphism:Itaffectsalargeproportionofthegenomeandhasavarietyofphenotypicconsequencesrelevanttohealthanddisease.Inspiteofthis,humangenomestructurevariationisincompletelycharacterizedduetoalackofapproachesfordiscoveringabroadrangeofstructuralvariantsinaglobal,comprehensivefashion.WeaddressedthisgapwithOpticalMapping,ahigh-throughput,high-resolutionsingle-moleculesystemforstudyinggenomestructure.WeusedOpticalMappingtocreategenome-widerestrictionmapsofacompletehydatidiformmoleandthreelymphoblast-derivedcelllines,andwevalidatedtheapproachbydemonstratingastrongconcordancewithexistingmethods.WealsodescribethousandsofnewvariantswithsizesrangingfromkbtoMb.structuralvariationcopynumbervariationopticalmappingsingle-moleculegenomicsgenomeassemblyRecentreports(1–11)havefirmlyestablishedgenomestructuralvariationasanimportantandpervasivesourceofgeneticpolymorphism.Sincetheinitialreports(1,2)ofwidespreadcopy-numbervariationbetweenthegenomesofphenotypicallynormalindividuals,investigatorshaveappliedhybridization-basedmethods(3,7,9,11),computationalapproaches(5,6),clonepaired-endsequencing(4,10)andmostrecentlyapaired-endsequencingbysynthesisapproach(8)tothediscoveryandcharacterizationofstructuralpolymorphism.Othershavedescribedphenotypicconsequencesofthesevariants,includingassociationswithmyocardialinfarction,neuroblastoma,autism,andschizophrenia(reviewedrecentlyinref. 12).Finally,theirconsistentassociationwithsegmentalduplicationsandotherclassesofrepeats(13)providesamechanisticexplanationfortheirorigin(14)andpointstoapreviouslyunappreciatedroleinevolution(15)aswellasdisease.Unfortunately,despiteallefforts,acomprehensivepictureofgenomestructurepolymorphismhasnotyetemerged.Currentgenome-widestudiesofstructuralvariationmanifestonlymodestconcordance,possiblyduetoascertainmentbiasesarisingfromthetechniquesemployed.Forexample,hybridization-basedmethods(2,3,7,9,11,16)aresubjecttononspecifichybridizationinrepeat-richregions,whileclone-basedstrategies(4,8,10)arelimitedbymaximumcloneinsertsizesandawideclonesizedistributionrelativetotheeventstheyaretryingtodetect.Morerecently,severalentirehumangenomesweresequencedusinghigh-throughputmethods(17–20),butthedifficultyofinterrogatingrepeat-richregionsiscompoundedbythesesystems’shortreadlengths.Inanefforttoovercomethesechallenges,wehaveappliedOpticalMappingtotheproblemofdiscerningstructuralvariationinnormalhumangenomes.OpticalMapping(21–35)isahigh-throughputsystemthatcombinessingle-moleculemeasurementswithdedicatedcomputationalanalysistoproduceorderedrestrictionmapsfromindividualmoleculesofgenomicDNA:essentially,asingle-moleculerealizationoftraditionalrestrictionfragmentlengthpolymorphismmapping(36).Eachsingle-moleculerestrictionmapisadirectmeasurementofthesourcegenome,freefrombiasesintroducedbycloning,amplification,orhybridization.Recentadvancesinsurfacechemistry,microfluidics,instrumentation,andalgorithms(SIText)haveincreasedoursystem’sthroughputsothatOpticalMappingisnowaviableplatformfortheanalysisofcomplexeukaryoticgenomes,includingthehumangenome.ThisreportpresentstheanalysisofstructuralvariationinfourhumangenomesusingOpticalMapping,comparestheseresultstoothergenome-wideanalysesanddescribesthousandsofpreviouslyunreportedstructuralvariants.ResultsOpticalMapConstruction.WeusedOpticalMapping(21–35andFig. 1)togenerateshotgunsingle-moleculerestrictionmapsfromthegenomesofacompletehydatidiformmole(37)(CHM1hTERT)andthreelymphoblast-derivedcelllines(GM15510,GM10860,GM18994).Highmolecular-weightgenomicDNAwasextractedfromthecellswithagentleliquidlysis,thendepositedonchargedglasssurfacesbyanarrayofmicrofluidiccapillarychannels(26).TheimmobilizedDNAmoleculesweredigestedinsituwiththemethylation-insensitiverestrictionendonucleaseSwaI,chosenbecauseitsmoderateaveragerestrictionfragmentsizebalancesgoodrestrictionmapresolutionwithaccuratefragmentsizing.ThedigestedDNAwasstainedwiththefluorescentdyeYOYO-1andimagedonalaser-illuminatedepifluorescencemicroscopyworkstationbuiltfromoff-the-shelfcomponents.Custommachine-visionsoftwareanalyzedthemicrographstoidentifythecleavedDNAfragments,estimatetheirsizesfromtheirfluorescence,andordercollinearfragments,producingorderedrestrictionmapsfromsinglemoleculesofDNA.Downloadfigure Openinnewtab Downloadpowerpoint Fig.1.AnoverviewoftheOpticalMappingplatform.Bulkmicroscopecoverglassiscleanedwithastrongacid,thentreatedwithasilanemixturetomakepositivelychargedOpticalMappingsurfaces(i).Asiliconwaferispatternedwithstandardphotolithographytechniques,andthenreplicatedintoaflexiblePDMSmicrofluidicdevice(ii)usingsoftlithography.Finally,pure,highmolecular-weightDNA(iii)isisolatedfromculturedeukaryoticcellsusingagentledetergent-basedlysisprotocol.ThemicrofluidicdeviceisadheredtotheOpticalMappingsurface,andtheDNAsolutionispumpedthroughthemicrochannels,whereintheDNAiselongatedandattachedtotheOpticalMappingsurfaceviaelectrostaticinteraction(iv).TheDNAisincubatedwitharestrictionendonuclease(v),whichcleavestheDNAatitscognatesites.ThecleavedDNAisstainedandimagedonanepifluorescencemicroscope(vi)illuminatedbyanargon-ionlaser(vii)andcontrolledbyacomputerworkstation(viii).AtightintegrationbetweencomponentsisresponsibleforOpticalMapping’shighthroughput.ThemicrofluidicdeviceconfinesDNAdepositiontoaregulargeometry,obviatingmanualmicroscopyandallowingasinglemicroscopetorunfor24 hunattended.LaserilluminationandasensitiveCCDcameraleverageYOYO-1’shighquantumefficiency,reducingper-imageexposuretimefromsecondstotensofmilliseconds.Finally,depositingthegenomicDNAwithcapillaryfloworientsallthemoleculesinthesamedirection,facilitatingreliablemachinevision.Thesesynergiesyieldathroughputof50,000–100,000moleculesanalyzedevery24 h,allowingdatacollectionfor50-foldcoverageofahumangenometobecompletedonasinglemicroscopeinaboutamonth.Theseshotgunsingle-moleculerestrictionmapsareassembledintogenome-wideconsensusrestrictionmapsusinganiterativeprocessinspiredbywhole-genomesequenceassembly(Fig. 2).Eachiterationhastwosteps,clusteringandassembly:Theclusteringstepgroupstogethersimilarsingle-moleculemapsbyaligningthemtoareferencemap(28,38),andthentheseclustersareassembledintoanewhypothesismapusingaBayesianmaximum-likelihoodassembler(39).ThefirstiterationusesareferencemapderivedinsilicofromtheNationalCenterforBiotechnologyInformation(NCBI)build35humanreferencesequence(40),butsubsequentiterationsofclusteringandassemblyextendandrefinethehypothesissothatthefinalconsensusmapsareanaccuraterepresentationofthegenomebeinganalyzed.Parametersforboththealignmentandassemblystepsaretunedsothatonlyhigh-qualitysingle-moleculemapsarepresentinthefinalassembly.Aftereightiterationsofassembly,thegenome-wideconsensusmapsthusconstructedspanasmuchas98.6%ofthegenomeandhaveanaverageassemblydepthofupto58-fold(Table 1andTable S1).Downloadfigure Openinnewtab Downloadpowerpoint Fig.2.Anoverviewofthemapassemblypipeline.ReferencemapsaregeneratedinsilicofromtheNCBIBuild35humangenomereferencesequence(40),andusedtoseedaniterativeprocessofpairwisealignment(whichclusterstogethersimilarsingle-moleculemaps)andlocalassembly(whichgeneratesaconsensusopticalmapfromaclusterofsingle-moleculemaps).Afterseveraliterationsofalignmentandassembly,theconsensusmapsarealignedbacktothereferencemapandanalyzedforplaceswheretheconsensusmapdifferssignificantlyfromthereference,indicatingpotentialpolymorphisms.Viewthistable:Viewinline Viewpopup Table1.OpticalmapcollectionandassemblystatisticsThegenome-wideconsensusmapsarehighlyaccurate:Inallfourgenomes,over95%ofthefragmentssize10 kbandgreaterarewithin10%oftheircorrespondingreferencefragmentsize.(Thisregime’sfragmentsizingerrorincreaseswithfragments 0.05)variantswereremoved,andtheremainingdifferenceswerefilteredconservativelytoremoveseveralclassesthat,basedonpastexperience,arelesslikelytorepresentbonafidesequencevariants.Afinalmanualcurationstepresolvedregionsthatwerenotamenabletoautomatedanalysis.DetailsregardingthesefilteringandcurationstepscanbefoundinSIText.Downloadfigure Openinnewtab Downloadpowerpoint Fig.3.ArepresentationofthestructuralvariationfoundinfourgenomesanalyzedbyOpticalMapping.VariantsfromtheCHMgenomearedepictedingreen;GM15510inblue;GM10860inred;andGM18994ingray.TheinsetdepictsfiveexampledifferencesfromthegenomeofGM10860:anextracut,amissingcut,a250 kbdeletion,a150 kbinsertion,anda150 kbinversion.WesummarizeourfindingsinTable 2.Thevariantsincludesimpleeventssuchasextrarestrictionsites,missingrestrictionsites,andinsertionsanddeletionswithsizedifferencesrangingfrommegabasesdownto3 kb.Theyalsoincludemorecomplexeventssuchasinversionsandlargediscordantregions.Intotal,wediscerned4,205uniquevariantsinthefourgenomesweanalyzed.WenotethatthesmallertotalfromCHMislikelyduetothereducednumberofsingle-moleculerestrictionmapscollectedfromthelimitedsampleavailable,resultinginalossofstatisticalpower.Wehypothesize,however,thatthislowerpowerissomewhatcompensatedforbythefactthatacompletehydatidiformmoleiseffectivelymonoploid(37),eliminatingtheeffectofdiploidyonanassemblerthatwasn’tdesignedtoaccommodatemixedhaplotypes.Viewthistable:Viewinline Viewpopup Table2.SummaryofstructuralvariantsdiscernedbyOpticalMappingWealsointersectedthevariantsfromeachgenomewithvariantsofthesametypefromtheotherthreegenomes(Table 2).Wenotethatoverathirdofthevariantswereportwereobservedinmultiplegenomes,givingusconfidencethattheseresultsareduetopolymorphismandnotthespuriousresultofcellcultureartifactsorotherrandomprocesses.[Theinfrequencyofculture-inducedartifactsisalsosupportedbyanalysesoftheHapMapparent-progenytrios(9).]Wesuggestthatthe322variantscommontoallfourgenomesmightbeduetoassemblyerrorsintheNCBIbuild35referencesequence,ortheymightrepresentpolymorphismsforwhichthereferencesequencereportsaminorfrequencyallele.ComparisontoOtherPlatforms.TovalidatethevariantsdiscernedbyOpticalMapping,wecarefullycomparedthemtoresultsreportedbyotherinvestigatorswhouseddifferenttechnologiestoanalyzesomeofthesamesamples(Table 3).Thereferenceplatform’sresultswerefilteredtoremovevariantsnotamenabletodetectionbyOpticalMapping(e.g.,inversionsthatwerecontainedentirelywithinasingleSwaIrestrictionfragment),andtheremainingvariantswerecomparedtotheconsensusmap.Table 3givesanoverviewofthesecomparisons,alongwiththeintersectionsofothertechnologies’results;notesoneachvariant’scomparisontotheopticalconsensusmapareincludedinTable S4,andadetailedexamplecomparingseveralfosmidend-sequencing(FES)andpaired-endmapping(PEM)variantstothecorrespondingopticalmapispresentedinFig. S2.Viewthistable:Viewinline Viewpopup Table3.SummaryofOMresultscomparedtootherplatformsBecauseFESandPEMtechnologieshavetheabilitytoestimateinsertionanddeletionsizesindependentofprobeplacementordensity,wealsocomparedthesizesofvariantsdiscernedwiththesetechnologiestothecorrespondingOpticalMappingvariants.Toincreasethelikelihoodthatthefindingsfromeachdatasetrepresentthesamesequence-levelevent,weonlyincludedOpticalMappingresultsthatmatchedone-to-onewithanFES-orPEM-derivedobservation.Wewereleftwith84pairsofobservationsforFESand82forPEM,severalofwhichwerediscardedaftermanualcuration(e.g.,toremoveseveralthatoverlappedgapsorwerepartsoflarge-scalediscordances).AlinearmodelfittotheremainingpairshasanR2of0.95andaslopeof0.97forFES,andanR2of0.94andaslopeof0.98forPEM,indicatingstrongagreementbetweenthesetwomethodsandOpticalMapping(Fig. S3andFig. S4).OpticalMappingComplementsOtherPlatforms.Aswewereperformingthecomparisondetailedabove,wenotedanumberofcommoncaseswhereOpticalMappingcomplementstheresultsofanotherplatform.Aparticularlystrikingexampleinvolveslargegainsinsequencediscernedbyhybridization-basedplatforms:suchresultscanindicateadditionalcopiesofasequence,butgivenoinsightintothegenomestructurethatengendersthegaininsequence.OpticalMapping’sabilitytoresolvestructuraldetailscanbringclaritytothissituation,asexemplifiedbyFig. 4:TheAffymetrix6.0SNPoligonucleotidemicroarrayindicateda290 kbgaininsequenceonGM10860chromosome16,andtheopticalmapidentifiesthiseventasaninvertedtandemduplication.Downloadfigure Openinnewtab Downloadpowerpoint Fig.4.Theopticalmapcomplementshybridization-basedapproaches.TheopticalmaprevealsthatthegaininsequencedetectedbytheAffymetrixSNP6.0platform(shadedregion)isduetoaninvertedtandemduplicationatthislocus(redarrows).End-sequencingstrategies,ontheotherhand,arelimitedintheirabilitytoresolvesequenceinsertionslargerthantheirinsert-orfragment-size,whileOpticalMappingissubjecttonosuchconstraints.Forexample,FESanalysisreportedbyKiddetal.(10)demonstratedthatclustersoffosmidswithonlyonealignedendcanindicatethepresenceofaninsertionthatwastoolargetobecapturedbythefosmidlibrary.OftheelevenclustersidentifiedbyKiddetal.,eighthaveclearsupportintheopticalmapandaninthcomesfromaregionoflargediscordancebetweentheopticalmapandthereferencegenome,makingthepresenceofextrasequencelikely(Table S4).(Severalhavesincebeenspannedbysequenceandcloselyagreewiththeopticalmap-derivedestimate.)Adetailedexample,includingmicrographsofsomeoftheDNAmoleculesthatsupportthisconclusion,ispresentedinFig. 5.Wealsofindevidencethatfosmidswithonealignedendthatoccuroutsideofclustersmightindicatesmallerinsertions:Aninterval-intersectionpermutationtest(seeSITextfordetails)revealsasignificantintersectionwithopticalmap-discernedinsertions(p  0.05)basedonanappropriatestatisticaltestoftheunderlyingsingle-moleculemapfragments.WealsoappliedasetofempiricallyderivedfilterstoaccountforothersourcesoferrorintheOpticalMappingprocess.Foradditionaldetail,seeSIText.Afinalmanualcurationstepservedtoelucidatehard-to-automatevariantssuchaslargeinversions.AcknowledgmentsWethankAlexLimforhispreliminarydata,andJonathanPritchardandStevenMcCarrollforstimulatingconversationsandearlyaccesstodata.ThisworkwassupportedbyNationalInstitutesofHealthGrantsR01HG000225andR33CA111933(D.C.S.),NationalResearchServiceAwardT32GM008349(S.R.),NationalResearchServiceAwardT32GM07215(B.T.),andNationalLibraryofMedicineTrainingGrant5T15LM007359(B.T.).Footnotes1Towhomcorrespondenceshouldbeaddressed.E-mail:dcschwartz{at}wisc.edu.Authorcontributions:B.T.,M.S.W.,S.G.,S.R.,D.S.,A.V.,andD.C.S.designedresearch;B.T.,K.P.,S.R.,C.L.,andM.K.-F.performedresearch;B.T.,M.S.W.,S.G.,K.P.,D.S.,A.V.,C.C.,J.M.K.,S.K.,R.R.,D.F.,M.A.N.,E.E.E.,M.K.-F.,U.S.,andM.L.contributednewreagents/analytictools;B.T.,M.S.W.,S.G.,K.P.,S.Z.,S.R.,D.S.,A.V.,C.C.,J.M.K.,M.A.N.,E.E.E.,andD.C.S.analyzeddata;andB.T.,S.G.,S.R.,andD.C.S.wrotethepaper.Theauthorsdeclarenoconflictofinterest.*ThisDirectSubmissionarticlehadaprearrangededitor.Thisarticlecontainssupportinginformationonlineatwww.pnas.org/lookup/suppl/doi:10.1073/pnas.0914638107/-/DCSupplemental.FreelyavailableonlinethroughthePNASopenaccessoption. References↵LucitoR,etal.(2003)Representationaloligonucleotidemicroarrayanalysis:Ahigh-resolutionmethodtodetectgenomecopynumbervariation.GenomeRes13:2291–2305.OpenUrlAbstract/FREEFullText↵SebatJ,etal.(2004)Large-scalecopynumberpolymorphisminthehumangenome.Science305:525–528.OpenUrlAbstract/FREEFullText↵SharpAJ,etal.(2005)Segmentalduplicationsandcopy-numbervariationinthehumangenome.AmJHumGenet77:78–88.OpenUrlCrossRefPubMed↵TuzunE,etal.(2005)Fine-scalestructuralvariationofthehumangenome.NatGenet37:727–732.OpenUrlCrossRefPubMed↵ConradDF,AndrewsTD,CarterNP,HurlesME,PritchardJK(2006)Ahigh-resolutionsurveyofdeletionpolymorphisminthehumangenome.NatGenet38:75–81.OpenUrlCrossRefPubMed↵McCarrollSA,etal.(2006)Commondeletionpolymorphismsinthehumangenome.NatGenet38:86–92.OpenUrlPubMed↵RedonR,etal.(2006)Globalvariationincopynumberinthehumangenome.Nature444:444–454.OpenUrlCrossRefPubMed↵KorbelJO,etal.(2007)Paired-endmappingrevealsextensivestructuralvariationinthehumangenome.Science318:420–426.OpenUrlAbstract/FREEFullText↵McCarrollSA,etal.(2008)Integrateddetectionandpopulation-geneticanalysisofSNPsandcopynumbervariation.NatGenet40:1166–1174.OpenUrlCrossRefPubMed↵KiddJM,etal.(2008)Mappingandsequencingofstructuralvariationfromeighthumangenomes.Nature453:56–64.OpenUrlCrossRefPubMed↵ConradDF,etal.(2010)Originsandfunctionalimpactofcopynumbervariationinthehumangenome.Nature464:704–712.OpenUrlCrossRefPubMed↵ZhangF,GuW,HurlesME,LupskiJR(2009)Copynumbervariationinhumanhealth,disease,andevolution.AnnuRevGenomHumG10:451–481.OpenUrlCrossRef↵BaileyJA,EichlerEE(2006)Primatesegmentalduplications:Cruciblesofevolution,diversityanddisease.NatRevGenet7:552–564.OpenUrlPubMed↵vanOmmenGJB(2005)Frequencyofnewcopynumbervariationinhumans.NatGenet37:333–334.OpenUrlCrossRefPubMed↵PerryGH,etal.(2006)Hotspotsforcopynumbervariationinchimpanzeesandhumans.ProcNatlAcadSciUSA103:8006–8011.OpenUrlAbstract/FREEFullText↵ShenF,etal.(2008)Improveddetectionofglobalcopynumbervariationusinghighdensity,non-polymorphicoligonucleotideprobes.BMCGenet9:27.OpenUrlCrossRefPubMed↵LevyS,etal.(2007)Thediploidgenomesequenceofanindividualhuman.PLoSBiol5:e254.OpenUrlCrossRefPubMed↵WheelerDA,etal.(2008)ThecompletegenomeofanindividualbymassivelyparallelDNAsequencing.Nature452:872–876.OpenUrlCrossRefPubMed↵WangJ,etal.(2008)ThediploidgenomesequenceofanAsianindividual.Nature456:60–65.OpenUrlCrossRefPubMed↵PushkarevD,NeffNF,QuakeSR(2009)Single-moleculesequencingofanindividualhumangenome.NatBiotechnol27:847–852.OpenUrlCrossRefPubMed↵SchwartzDC,etal.(1993)OrderedrestrictionmapsofSaccharomycescerevisiaechromosomesconstructedbyopticalmapping.Science262:110–114.OpenUrlAbstract/FREEFullText↵JingJ,etal.(1998)Automatedhighresolutionopticalmappingusingarrayed,fluid-fixedDNAmolecules.ProcNatlAcadSciUSA95:8046–8051.OpenUrlAbstract/FREEFullText↵LaiZ,etal.(1999)AshotgunopticalmapoftheentirePlasmodiumfalciparumgenome.NatGenet23:309–313.OpenUrlCrossRefPubMed↵LinJ,etal.(1999)Whole-genomeshotgunopticalmappingofDeinococcusradiodurans.Science285:1558–1562.OpenUrlAbstract/FREEFullText↵ZhouS,etal.(2003)Whole-genomeshotgunopticalmappingofRhodobactersphaeroidesstrain2.4.1anditsuseforwhole-genomeshotgunsequenceassembly.GenomeRes13:2142–2151.OpenUrlAbstract/FREEFullText↵DimalantaET,etal.(2004)AmicrofluidicsystemforlargeDNAmoleculearrays.AnalChem76:5293–5301.OpenUrlPubMed↵ValouevA,ZhangY,SchwartzDC,WatermanMS(2006)Refinementofopticalmapassemblies.Bioinformatics22:1217–1224.OpenUrlAbstract/FREEFullText↵ValouevA,etal.(2006)Alignmentofopticalmaps.JComputBiol13:442–462.OpenUrlCrossRefPubMed↵ValouevA,SchwartzDC,ZhouS,WatermanMS(2006)AnalgorithmforassemblyoforderedrestrictionmapsfromsingleDNAmolecules.ProcNatlAcadSciUSA103(43):15770–15775.OpenUrlAbstract/FREEFullText↵ZodyMC,etal.(2006)DNAsequenceofhumanchromosome17andanalysisofrearrangementinthehumanlineage.Nature440:1045–1049.OpenUrlCrossRefPubMed↵MitchelsonKRZhouS,HerschlebJ,SchwartzDC(2007)inNewHighThroughputTechnologiesforDNASequencingandGenomics,edMitchelsonKR(Elsevier,Amsterdam),pp266–301.↵ZhouS,etal.(2007)Validationofricegenomesequencebyopticalmapping.BMCGenomics8:278.OpenUrlCrossRefPubMed↵AnanievGE,etal.(2008)OpticalmappingdiscernsgenomewideDNAmethylationprofiles.BMCMolBiol9:68.OpenUrlCrossRefPubMed↵ChurchDM,etal.(2009)Lineage-specificbiologyrevealedbyafinishedgenomeassemblyofthemouse.PLoSBiol7:e1000112.OpenUrlCrossRefPubMed↵ZhouS,etal.(2009)Asinglemoleculescaffoldforthemaizegenome.PLoSGenet5:e1000711.OpenUrlCrossRefPubMed↵BotsteinD,WhiteRL,SkolnickM,DavisRW(1980)Constructionofageneticlinkagemapinmanusingrestrictionfragmentlengthpolymorphisms.AmJHumGenet32:314–331.OpenUrlPubMed↵JacobsPA,WilsonCM,SprenkleJA,RosensheinNB,MigeonBR(1980)Mechanismoforiginofcompletehydatidiformmoles.Nature286:714–716.OpenUrlCrossRefPubMed↵SarkarD(2006)Ontheanalysisofopticalmappingdata.PhDthesis(UniversityofWisconsin,Madison,WI).↵AnantharamanT,MishraB,SchwartzDC(1999)Proceedingsofthe7thInternationalConferenceonIntelligentSystemsforMolecularBiology,Genomicsviaopticalmapping.III:ContiginggenomicDNA(AAAIPress,MenloPark,CA),pp18–27.↵InternationalHumanGenomeSequencingConsortium(2004)Finishingtheeuchromaticsequenceofthehumangenome.Nature431:931–945.OpenUrlCrossRefPubMed↵RheadB,etal.(2010)TheUCSCgenomebrowserdatabase:Update2010.NucleicAcidsRes38:D613–D619.OpenUrlAbstract/FREEFullText↵RamanathanA,etal.(2004)AnintegrativeapproachfortheopticalsequencingofsingleDNAmolecules.AnalBiochem330:227–241.OpenUrlCrossRefPubMed↵JoK,etal.(2007)Asingle-moleculebarcodingsystemusingnanoslitsforDNAanalysis.ProcNatlAcadSciUSA104:2673–2678.OpenUrlAbstract/FREEFullText↵JoK,SchrammTM,SchwartzDC(2009)Asingle-moleculebarcodingsystemusingnanoslitsforDNAanalysis:Nanocoding.MethodsMolBiol544:29–42.OpenUrlCrossRefPubMed PreviousNext Backtotop ArticleAlerts UserName* Password* Submit EmailArticle ThankyouforyourinterestinspreadingthewordonPNAS.NOTE:Weonlyrequestyouremailaddresssothatthepersonyouarerecommendingthepagetoknowsthatyouwantedthemtoseeit,andthatitisnotjunkmail.Wedonotcaptureanyemailaddress. YourEmail* YourName* SendTo* Entermultipleaddressesonseparatelinesorseparatethemwithcommas. Youaregoingtoemailthefollowing High-resolutionhumangenomestructurebysingle-moleculeanalysis MessageSubject (YourName)hassentyouamessagefromPNAS MessageBody (YourName)thoughtyouwouldliketoseethePNASwebsite. YourPersonalMessage CAPTCHAThisquestionisfortestingwhetherornotyouareahumanvisitorandtopreventautomatedspamsubmissions. SendMessage CitationTools High-resolutionhumangenomestructurebysingle-moleculeanalysis BrianTeague,MichaelS.Waterman,StevenGoldstein,KonstantinosPotamousis,ShiguoZhou,SusanReslewic,DeepayanSarkar,AntonValouev,ChristopherChuras,JeffreyM.Kidd,ScottKohn,RodneyRunnheim,CaseyLamers,DanForrest,MichaelA.Newton,EvanE.Eichler,MarijoKent-First,UrvashiSurti,MironLivny,DavidC.Schwartz ProceedingsoftheNationalAcademyofSciencesJun2010,107(24)10848-10853;DOI:10.1073/pnas.0914638107 CitationManagerFormats BibTeX Bookends EasyBib EndNote(tagged) EndNote8(xml) Medlars Mendeley Papers RefWorksTagged RefManager RIS Zotero RequestPermissions Share High-resolutionhumangenomestructurebysingle-moleculeanalysis BrianTeague,MichaelS.Waterman,StevenGoldstein,KonstantinosPotamousis,ShiguoZhou,SusanReslewic,DeepayanSarkar,AntonValouev,ChristopherChuras,JeffreyM.Kidd,ScottKohn,RodneyRunnheim,CaseyLamers,DanForrest,MichaelA.Newton,EvanE.Eichler,MarijoKent-First,UrvashiSurti,MironLivny,DavidC.Schwartz ProceedingsoftheNationalAcademyofSciencesJun2010,107(24)10848-10853;DOI:10.1073/pnas.0914638107 ShareThisArticle: Copy TweetWidget FacebookLike Mendeley ArticleClassifications BiologicalSciencesAppliedBiologicalSciences TableofContents Submit SignupforthePNASHighlightsnewslettertogetin-depthstoriesofsciencesenttoyourinboxtwiceamonth: SignupforArticleAlerts Signup Jumptosection ArticleAbstractResultsDiscussionMaterialsandMethodsAcknowledgmentsFootnotesReferencesFigures&SIInfo&MetricsPDF YouMayAlsobeInterestedin HumanexploitationofCaribbeansharks AstudyfindsthathumanactivityhasledtoaseveredecreaseinCaribbeansharkabundance. Imagecredit:SeanMattson(InternationalCenterforTropicalAgriculture,Cali,Colombia). Climateandwingcolorationindragonflies Astudysuggeststhatmaledragonflieswillevolvesmallermelaninwingpatchesby2070asglobalwarmingcontinues. Imagecredit:Pixabay/liggraphy. COVID-19andairpollutiondisparities Thelargestpandemic-relateddeclinesinnitrogendioxidepollutionoccurredintheleastWhitecensustractsintheUnitedStates,whichnonethelessfacedhigherlevelsduringthepandemicthanmostWhitecommunitiesbeforethepandemic. Imagecredit:Pixabay/SD-Pictures. CoreConcept:InthewakeofCOVID-19,decentralizedclinicaltrialsgetpopular Theapproachcanenablefaster,morediversestudyenrollments.Butuncontrolledenvironmentscouldalsopotentiallyleadtofaultydataandflawedconclusions. Imagecredit:Shutterstock/Andrey_Popov. Opinion:Towardinclusiveglobalgovernanceofhumangenomeediting Genomeeditingtechnologyhasgrowntooquickly,andstakeholdersinthedebatearetoodiverse,forcurrentapproachestoestablisharobustregulatoryregime. Imagecredit:Shutterstock/vchal. SimilarArticles