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Analysis_1

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Analysis_1

    Analysis

Availableonlineatwww.sciencedirect.corn

    ;,

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    ;糖萨ScienceDirect

    ;J.Genet.Genomics35(2008)687695

    ;JOURNAL0

    ;GENE?CSAND

    ;GENO?CS

    ;www.jgenetgenormcs.org

    ;AnalysisofthemeiosisintheF1hybridsofLongiflorumxAsiatic

    ;(LA)oflilies(Lilium)usinggenomicinsituhybridization

    ;ShujunZhou’,MunikoteS.Ramannab,RichardG.EVisserb,JaapM.vanTuylb

    ;InstituteofLandscape,CollegeofAgriculture&Biotechnology,ZhejiangUniversity,Hangzhou310029,China

    ;PlantBreedingLab,WageningenUniversity,P.0.Box386,6700AJ,Wageningen,TheNetherlands

    ;Receivedforpublication21June2008;revised14July2008;accepted3September2008 ;Abstract

    ;LongiflorumandAsiaticliliesofthegenusLiliumofthefamilyLiliaceaearetwoimportantgroupsofmodemlilycultivars.Oneofthe

    ;maintrendsoflilybreedingistorealizeintrogressionbetweenthesegroups.Withcutstylepollinationandembryorescue.distanthybrids

    ;betweenthetwogroupshavebeenobtained.However,theFlhybridsarehighlysterileorsomeofthemcouldproduceasmallnumberof

    ;2ngametes,andtheirBC1progeniesareusuallytriploids.DutchlilybreedershaveselectedmanycultivarsfromtheseBClprogenies

    ;basedontheirvariation.Itispresumablysuggestedthatsuchvariationcouldbecausedbyintergenomicrecombinationandabnormal

    ;meiosisduringgameteformationinFlhybridsofLongiflorumxAsiatic(LA)hybridsinLilium.Therefore,themeioticprocessoftenF1

    ;LAhybridswascytologicallyinvestigatedusinggenomicinsituhybridizationandtraditionalcytologicalmethodsinthepresentresearch.

    ;Theresultsshowedthat:atmetaphaseI.thehomoeologouschromosomepairingamongdifferentFIhybridsrangedfrom2.0to11.4bi

    ;valentsformedbyhomoeologouschromosomesperpollenmothercell(PMC),andveryfewmultivalents,andevenveryfewbivalents

    ;wereformedbytwochromosomeswiminonegenomeratherthanhomoeologouschromosomesinsomePMCs;atanaphaseI,al1biva

    ;Ientsweredisjoinedandmostunivalentsweredivided.Boththedisjoinedbivalents(half-bi

valents)andthedividedunivalents(sister

    ;chromatids)movedtotheoppositepoles,andthenformedtwogroupsofchromosomes;bec

    ausethetworesultinghalf-bivalentsretained

    ;theiraxesinthecellundisturbed,manycrossovertypes,includingsinglecrossovers,threest

    randdoublecrossovers,fourstranddouble

    ;crossovers.fourstrandtriplecrossovers,andfourstrandmultiplecrossoversbetweentheno

    nsisterchromatidsinthetetradsofbivalents, ;wereclearlyinfefredbyanalyzingthebreakpointsonthedisjoinedbivalents.Thepresentin

    vestigationnotonlyexplainedthereasonfor

    ;sterilityoftheF1LAhybridsandthevariationoftheirBC

    progenies,butalsoprovidedanewmethodtoanalyzecrossovertypesinother

    ;Flinterspecifichybridsaswel1.

    ;Keywords:Lilium;genomicinsituhybridization;abnormalmeiosis;crossover;2ngamete

    ;Introduction

    ;Lilyisaveryimportantkindofcutflowers.SOfar. ;about8.000cultivarshavebeenregistered.Theyareclas

    ;siftedintothreegroups,i…eLongiflorum,Asiatic,and

    ;Orientalhybrids.TheyoriginatedfromcrosseswithinSec

    ;tionLeucolirion,SinomartagonandArchelirion,respec

    ;tively.TheLongiflorumcultivarspossessdown-ward

    shapedflowerswithdistinctivefragrance, ;whitetrumpet

    ;andcanbeeasilycultivatedyear-round;Asiaticcultivars ;Correspondingauthor.Tel:+8657186971684

    ;E.mailaddress:zhou2007@~u.edu.cn

    ;haveawiderangeofcolors(orange,yellow,white,pink, ;red,purple,andsalmon)withearlytolateflowering,and ;someofthemareresistanttoFusariumandviruses;and ;Orientalcultivarspossesspink.whiteoryellowlarge ;fowerswithasweetstrongfragrance,andmostofthem ;areresistanttoBotrytis.Thoughitisverydimculttocross ;betweendifferentgroupsorsections,oneofthetrendsof ;lilybreedingistorealizeintrogressionbetweengroups. ;Withcutstylepollinationandembryorescue,distanthy

    ;bridsbetweengroupshavebeenobtained.However,theFl ;hybridsarehighlysterileorsomeofthemcouldproducea ;littleamountof2ngametes,

    ;usuallytfiploids(Limeta1.,

    ;i.e.,theirBC1progeniesare

    ;2001;BarbaGonzalezeta1.,

    ;

    ;688ShujunZhoueta1./JournalofGeneticsandGenomics35(2oos)687695

    ;2004,2005a,2005b).Interestingly,Dutchlilybreeders ;haveselectedmanycultivarsfromsomeBC1progenies ;basedonthevariations.Genomicsituhybridization

;fGISI4)showedthat2ngametesandintergenomicrecom

    ;binationplaythevitalrolesinthevariation(Zhoueta1. ;20081.

    ;AdiploidspecieshastWOsetsofhomologouschromo

    ;somes.Eachchlomosomeofonesetmaypairwithacot

    ;respondingoneoftheothersetduringmeiosis.Suchnol’

    ;malinclosesproducengametes.2ngamete,i.e.,”unre—

    ;ducedorsomaticgamete”.possessesthesamechromo—

    ;somentnnbersassomaticcells.Itiscausedbyabnormal ;meiosisduetounfavourableenvironmenta1factorsnatu

    ;rallyorunpairingofhomoeologouschromosomesinin

    ;terspecifichybrids.Aninterspecifichybridcontainstwo ;setsofchromosomesoriginatingfrombothparentalspe

    ;cies,respectively.Thechromosomesetsofthehybridare ;knownashomoeologouschromosomes.Thistermisused ;todistinguishtwosetsofhomologouschromosomes ;withinaspecies.Becausenotal1chromosomescouldpair, ;interspecifichybridsusuallyhaveabnorma1meiosisand ;mightcausef0rmationof2ngametes.Thedirectconse

    ;quenceof2ngametesgivesrisetopolyploids.Ramanna

    2003)pointedoutthat2ngametehasmuch ;andJacobsen

    ;moreadvantagesoverchromosomedoublingforpoly

    ;ploidyandintrogressionbreeding.

    ;Intergenomicrecombinationresultsfi’omcrossovel’oc—

    ;curringbetweennonsisterchromatidsinvolvedintwo ;pairedhomoeologouschromosomes.whichisca1led”bi—

    ;valent”.Thus.crossoverisanimportanteventduring

    ;meiosis.Numerousstudiesonc1’OSSOVeFeventshavebeen

    ;reportedin0n,:”satit’”(Harushimaeta1..1998).Zea

    ;川叭?fAndersoneta1.,2003),Solanumtubero~’lllll(Jonge—

    ;dijkandRamanna.1989).HoMeumvulgare(Pickering, ;I991).A1abidopsisthalianafCopenhavei’eta1.,2004)and

    ;otherplantsfsee1.evie~bvNilssoneta1..1993).Ineal’15

    ;repol’ts.countin~chiasnlataatdiakinesisOl’atmetaphaseI

    ;ofmeiosiswithconventiona1stainingwastheIllain ;methodfoi’evahlatingCl’ossovereventsandrecombination.

    ;Thismethodisbeingreplacedbvmolecularmarkertech

    ;niqtiesthrough?hichgeneticlinkagemapscallbemade ;Genomicinsituhybridization.aDo,,el’fulandmodel’11

    ;cvtogenetictoo1callbeallidea1techniquetojnvesti~zate ;honloeologouschl’OlllOSOlllepail’ingandel’ossoverevents

    ;ofthemeiosisi11djsLanthvbl’id:-;ofLiliumIthasbeenused

    ;todjstinguishdiffel’enteenomesinintel’specifichybl’ids

    ;fSchwal’zachereta1..1989;Anamthawat—Jonssoneta1..

;1990;Kuipel’seta1..1997:Khrustalevaeta1..1998;Jiet

    ;a1..2004).tojnvestigatehomoeologouschromosomebe

    ;havioratlneiosjsofdistanthybridsfStevensonela1.. ;1998:Kamstraeta1..1999).andtodetectintergenomic ;rec0n1binantchrOInos0Inesanddeterminethefateofalien ;chromosomesintheirbackcrossprogeniesf-lacobseneta1.. ;1995:Takahashieta1.,1997:Khrustalevaeta..2000;

    ;Wangeta1..2005).Thistechniquehasa1sobeensuccess

    ;ful1vusedinLiliumtodiscriminateparentalgenomesof ;theF1interspecifichybridsandtodetectintergenomicre

    ;combinantchromosomesintheBC1andBC,progenies ;(Karloveta1..1999;Liraeta1..2000,200l2003; ;BarbaGonzalezeta1.,2004,2005a2005b).Fromthese ;extensivecytologicalanalysesoftheparentalF1hybrids ;andBCprogeniesinLilium.thefollowingconclusions ;werederived:(1)intheF1hybrids,bothfirstdivisionres

    ;titutionfFDR)andindeterminatemeioticrestitution(IMR) ;canresultin2ngameteformation;(2)manyBClprogenies ;haveintergenomicrecombinantchromosomeswhichresult ;fromcrossovel’sbetweenhomoeologouschromosomes

    ;duringgameteformationintheFIhybrids.Howevel’.how

    ;thehomoeologouschl’olnosoltlesbehaviourandhow

    ;crossoversoccurintheF1interspecifichybridsinLilium ;hasseldombeenstudied.

    ;Inthepresentarticle.10F1hybridswereinvestigated ;f0rtheirhomoeologouschromosomebehaviourduring ;micrOsDorogenesiswithconventionalmethodsandthe ;GISHtechnique.Theobiectivesweretoanalyzeho

    ;moeologouschromosomebehaviourandcrossoverevents ;andtoanswerthefollowingquestions.whichareimpor

    ;tantinlilybreeding:”WhyayelilyFIinterspec’(tich3’blids

    ;usualh’~terile?7z,,theyusuallyproducefz,,c?tionaf2n

    ;gametes?Isitpos’siblethatSOltlC()fthemcouldproduce

    ;notionalngamete?”

    ;Materialsandmethods

    ;Plantmaterials

    ;TendiploidFlinterspecifichybrids(2n=2x=24)uere ;usedtoinvestieatethehO111OeO10Ouschromosomebe

    ;havioul’duringmicrospol’ogenesisfTable11.Thesehybrids

    ;originatedfrOInC1’ossesberneenLongiflol’UIllcultivars

    ;fL-genome)andAsiaticcultivarsfA~enomeJ.whichbe

    ;1ongtoLeuco?rionsectional1dSinonlal’tagonsectionji1

    ;the~enusLilittm.1’espe’ctixel\AllthehybzidsweIeSLip

    ;pliedbythreeDutch1i1,,companies.i.eDeJongLelies

;BV.Tcstcentl’klnlBVandVletter&DenHaanBV.They

    ;ere~l’OWlli11greenhouseI)IlO’in(standardg1)?i11g

    ;condjLionsapp1icablerli1\cu1tivation.andarenlain

    ;tainedatPlantReseal’cl1Intel’nationa1.Wagenii1genU11i—

    ands. ;vel’sitx&Rcseal’chCentre.theNethel’

    ;Twodiploid?Ycultiva~’sf2n=2x=241.Asiatic’Mont

    ;Bant.andLoneiflorum.WhiteFox’.w’el’cusedtoextract

    ;genomicDNAinthjsresearch.’WhiteFox’DNA

    ;(Lgenome1wasusedasprobeandMonlBant’DNA

    ;(Agenome)asblockinGISH.

    ;Chromo~onlcprepatationwithanthers

    ;A1himpal’tofanthel’atmetaphaseIstagewascutand

    ;putonaslide.Underananatomicalmicroscope.the ;1.ab1e1

    ;ShujunZhoueta1./JournalofGeneticsandGenomics35(2008)687-695

    ;ChromosomepairingatmetaphaseIofmeiosesin10FjLAhybrids ;unnecessarydebrisofantherwal1wasremovedandthe ;pollenmothercellsfPMCs1werespreadontheslide,16 ;uLacetocarminwasswiftlyaddedandgentlymixedwith ;PMCs.Then,theslidewascoveredwithasquareclover ;glass,pressedgentlywiththethumb,andfinallyexamined ;withalightmicroscope.Goodslidesweresealedwithnail ;polisharoundcoverglassmargininordertoobserveand ;estimatetheextentanddifferenceofhomoeologouschro

    ;mosomepairing.Theaveragenumberofbivalentswasthe ;sumofbivalentsOfaUobservedPMCsdividedbythe ;numberofallobservedPMCs.

    ;Genomicinsituhybridization

    ;ChromosomepreparationwithanthersforGISHwas ;slightlydifferentfromthatusedfortheconventiona1cyto

    ;1ogica1method.Acetocarminwasreplacedbv45%acetic ;acid.Theslidewasdippedinliquidnitrogenforabout20s ;aftersquashing.Thecoverglasswasthenremovedwitha ;razorblade.Theslidewasputinpureethanolfor2rain, ;airdried.andstoredat-20.Cunti1use.

    ;110ta1genomicDNAwasextractedfromtheLongiflo