ARTICLEReceived20Dec2013|Accepted8Apr2014|Published8May2014Thetobaccogenomesequenceanditscomparison

withthoseoftomatoandpotato

NicolasSierro1,JamesN.D.Battey1,SoniaOuadi1,NicolasBakaher1,LucienBovet1,AdrianWillig1,w,SimonGoepfert1,ManuelC.Peitsch1&NikolaiV.Ivanov1

TheallotetraploidplantNicotianatabacum(commontobacco)isamajorcropspeciesandamodelorganism,forwhichonlyveryfragmentedgenomicsequencesarecurrentlyavailable.Herewereporthigh-qualitydraftgenomesforthreemaintobaccovarieties.ThesegenomesshowboththelowdivergenceoftobaccofromitsancestorsandmicrosyntenywithotherSolanaceaespecies.Weidentifyover90,000genemodelsanddeterminetheancestraloriginoftobaccomosaicvirusandpotyvirusdiseaseresistanceintobacco.WeanticipatethatthedraftgenomeswillstrengthentheuseofN.tabacumasaversatilemodelorganismforfunctionalgenomicsandbiotechnologyapplications.

OPEN

1PhilipMorrisInternationalR&D,PhilipMorrisProductsS.A.,2000Neuchatel,Switzerland.wPresentaddress:25bQuaiCharles-Page,CH-1205Gene`ve,

Switzerland.CorrespondenceandrequestsformaterialsshouldbeaddressedtoN.V.I.(email:Nikolai.Ivanov@pmi.com).

NATURECOMMUNICATIONS|5:3833|DOI:10.1038/ncomms4833|www.nature.com/naturecommunications1&2014MacmillanPublishersLimited.Allrightsreserved.Commontobacco(Nicotianatabacum)isoneofthemost

widelycultivatednon-foodcropsworldwideandisgrowninB120countries1.ItbelongstotheNicotianagenus,whichisnamedafterJeanNicotdeVillemainwho,in1560,becamethefirstpersontoimporttheseplantsfromtheAmericastoEurope.ThetermNicotianawasoriginallyusedbyAdamLonitzertodescribetobaccoplantsin1630(ref.2)andin1788byCarlvonLinne´(Linnaeus)todesignatetheentiregenus3.Over75naturallyoccurringNicotianaspecies,including49nativetoAmericaand25nativetoAustralia4,havebeenclassifiedbyGoodspeed5andKnapp6.MostcommercialtobaccoscultivatedtodaybelongtothespeciesNicotianatabacumL.,forwhich41,600N.tabacumcultivatedvarieties(cultivars)arelistedintheNationalPlantGermplasmSystem7.ThethreemostcommonlyusedtobaccotypesareFlue-Cured(orVirginia),BurleyandOriental,whicharetraditionallygrownandharvestedunderdifferentagriculturalpractices8.Tobaccoisamodelplantorganismforstudyingfundamentalbiologicalprocesses9,andisthesourceoftheBY-2plantcellline,whichisakeytoolforplantmolecularresearch10.Itisalsousedasamodelforplantdiseasesusceptibility,whichitshareswithotherSolanaceaeplantsincludingpotato,tomatoandpepper.Diseasesaffectingtobaccoincludethetobaccomosaicvirus(TMV),thetobaccoveinmottlingvirus(TVMV),thetobaccoetchvirus(TEV),andthepotatovirusY(PVY);theTN90varietyoftobacco,whichwesequencedhere,isnotableinthatishasbeenbredtoresisttheseviralinfections.Considerableinteresthascentredonunderstandingtheorigin,organizationandevolutionoftheN.tabacumgenome.Tobaccostandsoutasacomplexallotetraploidwithalarge4.5Gbgenomewithsignificantproportion(470%)ofrepeats11,12.Asaspecies,N.tabacum(2n¼4x¼48)evolvedthroughtheinterspecifichybridizationoftheancestorsofNicotianasylvestris(2n¼24,maternaldonor)andNicotianatomentosiformis(2n¼24,paternaldonor)about200,000yearsago13.Becauseofitscomplexityandimportance,thetobaccogenomeisatargetfortheSOL-100sequencingproject14,whichaimstodecipherthegenomesofthemostimportantSolanaceaespecies.Thegenomesequencesofmodernvarietiesofancestralspecieswererecentlyreported15,andlimitedevidencesuggeststhatNicotianaotophoraisanalternativepaternaldonor16,17.Inthisreport,however,wedemonstratethatthisisunlikelybecauseofthehighersequenceidentityoftheN.tabacumT-genomewiththatofN.tomentosiformis.Weshowthechromosomalrearrangementsbetweentheancestralandtobaccochromosomes,andprovideanexplanationforanapparentgenomereductionfollowingthehybridization.Inaddition,wepresentagenomiccomparisonoftobaccototwoothersolanaceousspecies,tomatoandpotato.Significantchromosomalreshufflingisclearlyobservedforallchromosomesdespitetheconservationoftheiroverallcount,confirmingpreviousreports18.Tobacco’srichmetabolism(involving44,000chemicalcomponents)andexceptionalabilitytoexpressproteins(440%ofitsdryweight)havepromptednumerousinitiativestoharnessitspotentialfortheproductionofbiologicallyactivesubstances19.

Here,wedescribethemajoralkaloidbiosynthesispathwayinNicotianaspecies,aswellasglutamate/aspartatepathwaysinthethreemaintobaccotypes.Inthiswork,wesequencethegenomesofkeyrepresentativesofthethreemajortypesoftobaccoandcombinethemwithgeneticandphysicalmapsoftobacco20,21.Weverifygenome

assemblyaccuracybymappingtranscriptomicsandExonArray22

dataofcorrespondingvarieties,weassesstheconsistencyofassembliesandpublishedphysicalandgeneticmaps,andwecompareN.tabacumS-andT-genomeswiththoseofN.sylvestrisandN.tomentosiformis.