ÿWPCo, À¢q×… p€>Éè„ ÄêH”`Ào¸É%×ÊQ”6ècÎ%‚h`ìâV—è&ˆgăr݇õiö–³Ï»%)V=ÏCI_ÞM³\'ÀFÈ,º“I¼èø,5éIVäu÷ÓÁrp@§hZôæ¹1‚= rnÃ>Óâ¡@TOi9q¡£mïÏ8˜uPØ’SUèÔ3š@™O^Ã_r¡€·4HžiÜÝáÕVù$ŠÎéYÑȦS˜%‡æñŽªëª…eËãÒ~PNÿRÝRù߉ÏרÉÎ+Ëš»Õ>:«/²¿pê}ÑÏõäô…e‰)·Ï 2«1‡k×? °³TOÉ ˆ7ÖOIÖ‡îˆ78ßæ%>!@‡5ÖHŽ>s_|T —Ñùø£Ã޲À 4A/]û#—³¹¯ÂºE4A&¼Íç5AûÈcqð É “ŸäG©•›a™d/Ų>ÌÉ/GÕÓc»yƒôÍÅs[9¡tð pébû³Y3¸Qù? 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òˆÌÌóóó ób* ÿÿ8Bullet ListIndented Bullet Listà0..àððà0` ..` àÌ<þ6X9`("Courier 10cpi-xÐðA`&UniversScalable,xÐðA`&UniversScalable*Ͼ¦ `(CG TimesScalable  ÿÿ3sy…‹“›¡1Right-Aligned Paragraph NumbersI.A.1.a.(1)(a)i)a)#|«<þ6X9`("Courier 12cpiôÂdþ6X@Ƀ8Ç;ô@<þ6X9`("Courier 10cpiXôxþ6X@É“8Ç;X@-xÐðA`&UniversScalable–– xÐðPö»7û–P,xÐðA`&UniversScalableddxÐðPö»7ûdP*Ͼ¦ `(CG TimesScalableXXwϾ¦ P쉊7ûXP d˜HP LaserJet 4,ü,,,,,ü0($¡¡ëU+&´/„ÿU‹ÿÀÀÀÝ ƒ7!ÝÝ  ÝÔ€XZXõôÔÔ€.XZXXXZÔò òÑ8€.Z.XXdðdÈ8ÑÓ=€.Z.XÐXdÐd=ÓÓÓÔ_Ô€€€Genetic€Mapping€of€Loci€Underlying€Field€Resistance€to€SoybeanÐ ° Ðà@\ \ œìàSudden€Death€Syndrome€(SDS).ˆÐ à Ðó óÌÌ€€€€€€€N.€Hnetkovsky,€S.J.C.€Chang,€T.W.€Doubler,Ì€€€€€€€€€€€P.T.€Gibson€and€D.A.€Lightfootòò*óó.Ð  p  ÐÌÌ€Dept.€of€Plant€and€Soil€Science,€Molecular€Science€Program,€€€€€€Ï€€€€€€€Southern€Illinois€Univ.,€Carbondale,€IL€62091.ÌÌÌÌAbbreviations:€DI,€disease€incidence;€DS,€disease€severity;€QTL,Ïquantitative€trait€loci;€RFLP,€restriction€fragment€lengthÏpolymorphism;€RAPD,€random€amplified€polymorphic€DNA;€RIL,Ïrecombinant€inbred€line.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€Ð à0" ÐóóÌS.J.C.€Chang,€T.W.€Doubler,€P.T.€Gibson€and€D.A.€Lightfootòòóó.€Dept.Ð #P& Ðof€Plant€and€Soil€Science,€Molecular€Science€Program,€SouthernÏIllinois€Univ.,€Carbondale,€IL€62091.€N.€Hnetkovsky,€OlneyÏCentral€College,€Olney€IL€62450.€Received€March€19,€1995.ÏAccepted€April€17,€1995.€*€Corresponding€author.€E„mailÏGA4082@SIUCVMB.SIU.EDU.Ð  Ð* &0 Ðò ò€€€€€€€€€€€€€€€€€€€€€€€€€ABSTRACTÌó óSudden€death€syndrome€(SDS)€€is€a€fungal€disease€of€soybeanÏ(òòGlycine€maxóó€(L.)€Merr.),€caused€by€òòFusarium€solanióó€(Mart.)€Sacc.€Ïf.€sp.€òòphaseolióó€(Burk.)€Snyd.€&€Hans.,€type€A,€that€reduces€cropÏyields€in€the€US.€Quantitative€partial€resistance€to€SDS€doesÏexist;€therefore€one€method€of€controlling€the€disease€is€to€selectÏcultivars€with€genetic€resistance.€€The€objective€of€this€study€wasÏto€identify€and€locate€alleles€of€chromosomal€segments€associatedÏwith€field€resistance€to€SDS€in€adapted€soybean€genotypes€usingÏmolecular€markers.€Seventy€polymorphic€DNA€markers€were€comparedÏwith€SDS€response€among€100€Fòò5:9óó€recombinant€inbred€lines€derivedÏfrom€a€cross€between€SDS€resistant€'Forrest'€and€SDS€susceptibleÏ'Essex'.€SDS€disease€incidence€(DI),€disease€severity€(DS)€andÏyield€were€determined€in€replicated,€SDS€infested€test€sites€duringÏfour€years€encompassing€five€locations€using€recombinant€inbredÏlines€from€the€Fòò5:7óó€to€Fòò5:11óó.€Two€separate€chromosomal€segmentsÏidentified€by€two€RAPD€markers,€OO05òò250óó€and€OC01òò650óó,€were€found€to€beÏassociated€with€mean€SDS€response€across€5€locations€as€well€asÏwithin€each€of€the€five€locations.€These€two€quantitative€traitÏloci€(QTL)€jointly€accounted€for€34€%€of€total€phenotypicÏvariability€in€mean€DI.€OC01òò650óó€was€significantly€associated€withÏmean€DS€and€yield€and€was€putatively€assigned€to€linkage€group€N.ÏThe€beneficial€allele€was€derived€from€the€resistant€parentÏForrest.€OO05òò250óó€was€not€significantly€associated€with€mean€DS€orÏyield€and€was€putatively€assigned€to€linkage€group€C.€TheÏbeneficial€allele€òòóówas€derived€from€the€susceptible€parent€Essex.€Molecular€markers€can€be€used€to€define€alleles€of€chromosomalÏsegments€conferring€resistance€to€SDS€in€several€environments€andÏmay€allow€efficient€selection€of€resistant€genotypes€with€goodÏyield€potential€for€SDS€infested€fields.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÇ€€€€€Soybean€sudden€death€syndrome€(SDS)€is€causing€increasinglyÏsevere€losses€to€soybean€(òòGlycine€maxóó€(L.)€Merr.)€productionÏ(Mulrooney€et€al.,€1988;€Gibson€et€al.,€1993;€Yang€and€Rizvi,Ï1994).€The€inciter€of€SDS€is€òòFusarium€solanióó€(Mart.)€Sacc.€f.€sp.Ïòòphaseolióó,€type€A€(Rupe,€1989;€Roy,€1989òòóó,€O'Donnell€and€Gray,€1995).ÏTo€date,€host€plant€resistance€has€been€the€most€effective€controlÏmeasure€for€SDS€(Gibson€et€al.,€1994).€However,€SDS€resistance€isÏdifficult€to€determine€since€resistance€is€incomplete€(partial),Ïpolygenic€in€nature€and€can€be€environmentally€sensitive.Ì€€€€€€Classical€genetic€inheritance€studies€in€the€field€estimatedÏthat€SDS€resistance€(expressed€as€disease€incidence€or€diseaseÏseverity)€is€conditioned€by€a€minimum€of€five€loci€in€each€of€twoÏprogeny€populations€of€adapted€soybean€(Matthews€et€al.,€1991;ÏNjiti€et€al.€1995).€In€a€third€population€a€genetic€study€using€aÏgreenhouse€assay€indicated€resistance€to€leaf€symptom€development,Ïexpressed€as€disease€severity,€may€be€conditioned€by€a€single€gene,Ïòòrfsóó€(Stephens€et€al.,€1993).€However,€in€the€field€òòrfsóó€confers€onlyÏpartial€SDS€resistance€(Gibson€et€al.,€1994).€Allelic€relationshipsÏamong€resistance€loci€in€the€three€separate€progeny€populations€areÏunknown.Ì€€€€€All€soybean€genotypes€tested€show€some€SDS€disease€symptoms€inÏinfested€fields€during€conditions€conducive€to€disease€expressionÏ(Gibson€et€al.,€1994).€Therefore,€resistance€to€SDS€in€soybeanÏgermplasm€is€unlikely€to€show€the€classical€race„cultivar„specificÏresistance€manifest€in€the€hypersensitive€response.€Rather,€usefulÏfield€resistance€appears€to€be€partial€€and€polygenic€(Matthews€et€al.,€1991;€Gibson€et€al.,€1994;€Njiti€et€al.,€1995).€€Ì€€€€€€€Breeding€for€SDS€resistance€is€complicated€by€the€absence€ofÏa€satisfactory€quantitative€greenhouse€assay€predictive€of€fieldÏperformance,€thereby€necessitating€extensive€field€selections€withÏmany€lines.€Factors€such€as€rainfall,€soil€fertility€,€maturityÏdate,€date€of€planting,€plant€genotype€and€cyst€nematodeÏinfestation€€all€significantly€alter€the€degree€of€diseaseÏprogression€in€the€field€(Rupe€et€al.,€1991;€Rupe€et€al.,€1994;ÏGibson€et€al.,€1994).€Therefore,€reliable€field€evaluations€requireÏmultiple€replications€in€many€infested€sites€with€accurate€diseaseÏprogression€data€collected€at€3„4€day€intervals€(Gibson€et€al.,Ï1994;€Njiti€et€al.,€1995).€Consequently,€conventional€breeding€forÏSDS€resistance€is€time€consuming,€difficult€and€expensive.Ì€€€€€€€€Molecular€biotechnology€makes€it€possible€to€identifyÏchromosomal€segments€containing€agriculturally€important€genes€asÏquantitative€trait€loci€(QTL)€using€DNA€markers.€DNA€markers€canÏexpedite€the€identification€of€SDS€resistant€soybean€lines€based€onÏgenotype€instead€of€phenotype,€which€should€reduce€the€need€forÏreplicated€field€tests.€Potentially,€lines€containing€severalÏresistance€genes€or€combinations€of€SDS€resistance€loci€withÏdifferent€modes€of€action€can€be€selected€with€molecular€markers.Ì€€€€€€€€€In€studying€disease€resistance€in€plants€the€use€of€DNAÏmarkers€has€focused€on€genes€involved€in€the€race„cultivar€specificÏhypersensitive€response€(complete€resistance).€These€include€singleÏgene€resistance€to€fungal€pathogens€in€tomato€(òòLycopersicumÏlycopersiconóó€L.)€(Sarfati€et€al.,€1989);€lettuce€(òòLactuca€sativaóó€L.)€(Michelmore€et€al.,€1991);€rice€(òòOryza€sativaóó€L.)€(Yu€et€al.,Ï1991);€and€soybean€(Diers€et€al.,€1992).Ì€€€€€€€€Using€DNA€markers,€cyst€nematode€resistance€has€been€shownÏto€be€a€race„cultivar€specific€partial€resistance€mediated€by€theÏcombined€activity€of€2„3€loci€in€potato€(òòSolanum€spegazziniióó€L.)Ï(Kreike€et€al.,€1994)€and€soybean€(Concibido€et€al.,€1994;€Webb€etÏal.,€1995).€However,€in€some€cases,€resistance€may€be€considered€aÏspecialized€example€of€race„cultivar€specific€complete€resistanceÏmediated€by€a€hypersensitive€response€(Endo€et€al.,€1964;€Webb€etÏal.,€1995).€Ì€€€€€€€€DNA€markers€have€not€been€widely€used€in€studying€partialÏdisease€resistances€which€are€based€on€interactions€between€severalÏloci€in€plants.€Those€reported€include€partial€resistance€to€fungalÏpathogens€in€corn€(òòZea€maysóó€L.)€(Bubeck€et€al.,€1993),€mung€beanÏ(òòVigna€radiataóó€L.)€(Young€et€al.,€1994),€rice€(Wang€et€al.,€1994)Ïand€barley€(òòHordeum€vulgaraeóó€L.)€(Backes€et€al.,€1995);€and€partialÏresistance€to€bacterial€pathogens€in€bean€(òòPhaseolus€vulgarisóó€L.)Ï(Nodari€et€al.,€1993)€and€tomato€(Danesh€et€al.,€1994;€Sandbrink€etÏal.,€1995).€However,€breeding€for€partial€resistance€to€fungalÏpathogens€is€relatively€common€because€of€the€stable€nature€of€theÏdisease€resistance€(Eddington€et€al.,€1994;€Gingera€et€al.,€1994)Ïand€because€genes€conferring€partial€disease€resistance€can€affordÏcross€protection€to€several€fungal€pathogens€(Zhang€et€al.,€1994).ÏCross€protection€is€also€manifest€in€systemic€acquired€resistanceÏ(Ward€et€al.,€1991)€and€is€mediated€in€part€by€genes€conferringÏpartial€resistance€to€fungal€pathogens€(Alexander€et€al.,€1993).€€Selection€for€partial€resistance€can€be€preferable€to€single€geneÏresistance€since€partial€resistance€may€be€stable€to€geneticallyÏvariable€field€populations€of€pathogens€(Eddington€et€al.,€1994),Ïincluding€òòF.€solanióó€f.€sp.€òòphaseolióó,€type€A.€DNA€marker€studiesÏindicate€that€some€partial€disease€resistance€loci€areÏenvironmentally€insensitive€in€the€field€and€may€be€raceÏinsensitive€€(Bubeck€et€al.€1992;€Wang€et€al.€1994).Ì€€€€€€€This€paper€demonstrates€the€utility€of€DNA€genetic€markersÏfor€identifying€chromosomal€segments€contributing€partialÏresistance€to€SDS€in€the€field.€The€implications€for€the€use€of€DNAÏmarkers€for€improved€selection€of€SDS€resistance€in€soybean€areÏconsidered.ÇÌ€€€€€€€€€€€€€€€€€€€€€ò òMATERIALS€AND€METHODSó óÌÌò òPlant€Materialó óÌ€€€€€The€'Essex'€by€'Forrest'€(ExF)€Fòò5óó€derived€òòóópopulation€of€soybeanÏrecombinant€inbred€lines€(RILs)€was€constructed€by€crossing€EssexÏ(Smith€and€Camper,€1973)€and€Forrest€(Hartwig€and€Epps,€1973).ÏEssex€is€SDS€susceptible,€while€Forrest€is€SDS€resistant€(Fig.€1;ÏGibson€et€al.,€1994).€About€4500€Fòò2óó€plants€were€inbred€to€the€Fòò5óóÏgeneration€using€a€single€pod€descent€method€(Brim,€1966).€In€1988,Ïa€random€bulk€of€seed€was€planted€to€obtain€500€Fòò5óó€plants€of€whichÏ150€were€randomly€selected,€only€intentionally€excluding€a€fewÏagronomically€undesirable€extremes.€In€1989,€100€Fòò5óó„derived€linesÏof€modal€maturity€(mid„maturity€group€V)€with€sufficient€seeds€forÏfield€testing€were€retained.€All€inbreeding€occurred€in€fields€withÏminimal€incidence€of€soybean€cyst€nematode€(SCN)€and€no€history€ofÏSDS.€Ìò òSDS€Disease€Scoringó óÌ€€€€Eight€field€experiments€were€conducted€over€a€four€year€periodÏencompassing€a€total€of€eight€locations€in€southern€Illinois,€ofÏthese€useful€data€could€be€derived€from€five€locations.€The€otherÏthree€were€excluded€due€to€poor€stand€and€absent€or€minimal€SDSÏleaf€symptoms€resulting€from€drought.€The€five€were€Villa€RidgeÏ1990€(V90);€Cora€1991€(C91);€Pulaski€1991€(P91);€Cora€1992€(C92);Ïand€Ridgway€1993€(R93)€(C91€and€C92€were€separate€fields,€2€kmÏapart).€All€fields€were€selected€based€on€a€history€of€visually€uniform€SDS€infestation€and€managed€as€described€by€Gibson€et€al.,Ï(1994).Ì€€€€€The€100€ExF€Fòò5óó€derived€lines€were€€scored€for€DI,€DS€and€yieldÏas€described€(Matthews€et€al.,1991;€Gibson€et€al.,€1994).€Two€rowÏplots€in€a€partially€balanced€simple€11€x€11€lattice€design€(2Ïreplications,€8€duplications€per€parent€and€5€checks)€were€usedÏ(Gomez€and€Gomez,€1984).€Disease€was€rated€weekly€and€the€lastÏscore€before€and€the€first€score€after€R6€(full€pod)€were€used€toÏstandardize€DI€and€DS€to€the€R6€stage.€DI€was€defined€as€theÏpercentage€of€plants€in€the€plot€with€visible€leaf€symptoms.€DS€wasÏrated€as€the€degree€of€leaf€damage€on€diseased€plants€and€wasÏscored€on€a€scale€of€1€to€9€(1€=€0„10%/1„5%,€2€=€10„20%/6„10%,€3€=Ï20„40%/10„20%,€4€=€40„60%/20„40%,€5€=€>60%/40%€chlorosis/necrosisÏwhereas€6€=€1„33%,€7€=€34„66%,€8€=€66„100%€premature€defoliationÏand€9€=€premature€death)(Gibson€et€al.€1994).€Yield€was€determinedÏat€harvest€from€the€two€row€plots€trimmed€to€3€m€(4.3€m€planted).ÏThere€was€no€intra„plot€bordering€since€the€progeny€populationÏdisplayed€uniform€growth€habit€and€maturity€dates.€Lattice€adjustedÏmeans€were€used€whenever€the€lattice€analysis€was€more€efficientÏthan€the€RCB.€To€detect€transgressive€segregants,€an€LSD€(òòPóó€=€0.05)Ïfor€across€environment€means€of€individual€lines€vs.€parents€wasÏcalculated€using€the€genotype€x€environment€interaction€as€theÏerror€variance.ÌÌò òDNA€Clonesó óÌ€€€€€Bacterial€strains€containing€cloned€soybean€òòPstIóó€genomic€DNA€inserts€were€obtained€from€Dr.€R.€Shoemaker,€USDA€ARS,€Ames,€IowaÏ(Shoemaker€and€Olson,€1993).ÌÌò òPlant€DNA€€Extraction,€Restriction€Digestion,€Electrophoresis€andÏSouthern€transfersó óÌ€€€€€€Leaf€and€seed€material€were€obtained€from€Dr.€Paul€Gibson€andÏDr.€Michael€Schmidt,€Southern€Illinois€University€at€Carbondale.€ÏLeaf€material€for€DNA€extraction€was€collected€from€20€Fòò5:9óó€fieldÏgrown€plants€per€genotype€in€July€1992€(Agronomy€Research€Center,ÏSouthern€Illinois€University€at€Carbondale)€and€frozen€(„70òòoóóC).Ì€€€€€DNA€extraction€from€leaf€tissue€was€according€to€Dellaporta€etÏal.,€(1983)€with€modification€for€legumes€(Guo€et€al.,€1991).ÏAliquots€of€soybean€DNA€were€digested€with€one€of€five€restrictionÏenzymes:€òòTaqóóI,€òòDraóóI,€òòEcoóóRI,€òòEcoóóRV,€or€òòHindóóIIIòòóó€based€on€informativeÏprobe€enzyme€combinations.€DNA€fragments€were€resolved€by€gelÏelectrophoresis€on€a€10€g€Lòò„1óó€agarose€gel,€then€transferred€andÏfixed€to€Hybond„N+€(Amersham€Corporation,€Arlington€Heights,€IL).€Ì€€€€€€The€informative€probe„enzyme€combinations€were€obtained€fromÏDr.€P.€Keim€(N.€Arizona€University,€Flagstaff,€AZ),€Dr.€R.ÏShoemaker€(USDA„ARS,€Ames,€IA),€Dr.€D.€Webb€(Pioneer€Hybrid,ÏJohnston,€IA)€or€were€generated€in€this€study.€In€total,€243€RFLPÏprobes€were€tested€for€the€ability€to€detect€polymorphism€betweenÏEssex€and€Forrest.€Polymorphic€RFLP€loci€were€referred€to€using€theÏnaming€convention€suggested€by€the€Soybean€Genetics€CommitteeÏ(Soybean€Genetics€Newsletter,€1995,€22:11„22).€Ìò òÌDNA€Hybridizationsó ó€Ì€€€€€Plasmid€DNA€was€digested€with€òòPstóóI€and€inserts€were€purifiedÏby€gel€electrophoresis€and€elution€for€use€in€radio„labelingÏreactions€as€described€by€Guo€et€al.€(1991).€Briefly,€50€to€100€ngÏof€probe€DNA€was€labelled€with€òò32óóP„dCTP€by€random€hexamer€primedÏsynthesis.€Hybridization€was€according€to€Guo€et€al.,€(1991)€exceptÏfor€the€following€modifications:€Filters€were€hybridized€at€65òòoóóC€inÏthe€rotator€incubator.€Polyethylene€glycol€8,000€at€5%€(w/v)€wasÏused€in€hybridization€media€in€place€of€5%€(w/v)€Dextran€SulfateÏ500,000.ÌÌò òRAPD€protocol„Polymerase€Chain€Reactionó óÌ€€€€The€amplification€reactions€were€performed€after€Williams€etÏal.€(1990)€with€180€separate€primers€from€kits€A,€B,€C,€D,€E,€F,€G,ÏH€and€O€from€Operon€Technologies€(Almeda,€CA).€The€total€volume€ofÏ25€ððl€€contained€€100€ððM€dATP,€dCTP,€dGTP,€and€dTTP,€0.2€ððM€of€a€10Ïbase€pair€primer,€10€„€40€ng€genomic€DNA,€and€either€1.0€unit€òòTaqóóÏpolymerase€with€1.5€mM€MgClòò2óó€(Promega,€Madison,€WI;€Perkin€ElmerÏCetus,€Norwalk,€CT)€€or€2.5€units€of€the€Stoffel€fragment€of€òòTaqóóÏpolymerase€with€3€mM€MgClòò2óó.€DNA€was€amplified€in€a€thermal€cyclerÏ(Savant,€New€York,€NY)€programmed€for€45€cycles€of€1€minute€at€94òòoóóC,Ï1€minute€at€36òòoóóC,€and€2€minutes€at€72òòoóóC.€Amplification€products€wereÏanalyzed€by€electrophoresis€on€a€14€g€L€òò„1óó€agarose€gel€stained€withÏethidium€bromide.€Primers€which€generated€distinct€polymorphicÏbands€between€Essex€and€Forrest€across€a€range€of€soybean€DNAÏconcentrations€(10€„€40€ng)€were€identified€and€used€to€amplify€DNA€samples€from€the€Fòò5:9óó€lines€to€generate€segregation€data€for€mappingÏanalysis.€RAPD€markers€amplified€with€Stoffel€fragment€reportedÏhere€were€OO05òò450óó€and€OO05òò250óó.€All€others€were€amplified€by€òòTaqóóÏpolymerase.€RAPD€markers€associated€with€SDS€resistance€wereÏamplified€independently€on€three€or€more€separate€occasions€toÏassure€reproducibility.€Polymorphic€RAPD€loci€were€referred€toÏusing€the€naming€convention€suggested€by€the€Soybean€GeneticsÏCommittee€(Soybean€Genetics€Newsletter,€1995,€22:11„22).ÌÌò òMapping€Quantitative€Resistance€Loció óÌ€€€€To€discover€genomic€regions€associated€with€SDS€resistance€theÏRILs€were€classified€as€Essex€type€or€Forrest€type€(heterozygotesÏwere€excluded)€for€òòóóeach€marker€and€compared€with€SDS€diseaseÏresponse€scores€by€analysis€of€variance€(ANOVA).€One„way€ANOVA€wasÏperformed€with€SAS€(SAS€Institute€Inc.,€Cary,€NC).€The€probabilityÏof€association€of€each€marker€with€each€trait€was€determined€and€aÏsignificant€association€was€declared€if€òòPóó€òò<óó€0.005€(unless€notedÏotherwise€in€the€text)€to€minimize€the€detection€of€falseÏassociations€(Lander€and€Botstein€1989).€Mapmaker„EXP€3.0€(LandersÏet€al.,€1987)€was€used€to€calculate€map€distances€(cM)€betweenÏlinked€markers€using€the€RIL€(ri„self)€genetic€model.€To€identifyÏintervals€containing€QTL€governing€SDS€response€the€marker€map€andÏdisease€data€were€simultaneously€analyzed€with€Mapmaker/QTL€1.1Ï(Paterson€et€al.,€1988)€using€the€Fòò2óó„backcross€model€for€traitÏsegregation€(Webb€et€al.,€1995,€M.J.Daly,€personal€communication,ÏWhitehead€Instit.,€Cambridge,€MA,€1995€).€Putative€QTL€were€inferred€when€LOD€(logòò10óó€of€odds€ratio)€scores€exceeded€2.0€at€someÏpoint€in€each€interval.ÌÇò òRESULTS€AND€DISCUSSIONó óÌò òÌPolymorphism€and€Linkage.ó óÌ€€€A€total€of€243€selected€DNA€clones€and€180€random€RAPD€primersÏwere€surveyed€for€polymorphism€between€Essex€and€Forrest.€Only€58ÏRFLP€probes€and€32€RAPD€primers€gave€clear€polymorphism.€SinceÏEssex€and€Forrest€are€more€closely€related€than€the€parents€of€mostÏmapping€populations€as€judged€by€their€co„variance€coefficientÏ(0.21)€(Allen€and€Bhardwaj,€1987)€and€RFLP€polymorphism€indexÏ(0.29)€(Keim€et€al.,€1992;€Skorupska€et€al.,€1994)€€the€low€RFLPÏfrequency€(0.24)€and€RAPD€frequency€(0.18)€was€expected.€€Twenty€ofÏthe€58€RFLP€markers€were€selected€as€likely€to€represent€separateÏloci€(>€10€cM€apart)€and€likely€to€identify€the€loci€mapped€inÏòòG.maxóó€x€òòG.sojaóó€(Shoemaker€and€Olsen,€1993).€These€20€RFLP€markersÏwere€scored€in€the€segregating€populationand€and€identified€22Ïloci.€The€32€RAPD€primers€were€scored€and€identified€48€loci.€InÏtotal€70€loci€were€mapped,€57€mapped€to€eight€coherent€linkageÏgroups€encompassing€1,065€cM.€There€were€13€unlinked€markers€whichÏtogether€with€the€linkage€groups€would€allow€detection€ofÏassociations€with€SDS€resistance€QTL€over€about€1,551€cM€with€meanÏintervals€of€òòóó18.7€cM.€This€compares€to€a€recombination€distance€forÏthe€soybean€genome€of€nearly€3,000€cM€within€25€linkage€groupsÏ(Shoemaker€and€Olson,€1993).€The€Fòò5:9óó€lines€were€heterogeneous€andÏthis€was€detected€with€RFLP€markers.€Heterogeneity€is€a€product€ofÏthe€heterozygosity€€present€in€each€Fòò5óó€plant€used€for€lineÏderivation.€Theoretically€the€progeny€line€mean€heterogeneity€would€be€6.25%€of€the€genome.€Mean€heterogeneity€was€calculated€from€dataÏfor€all€co„dominant€RFLP€probes€within€all€the€RILs€and€was€8.04%Ïwhich€was€not€significantly€different€from€the€6.25%€expected€(òòPóó€<Ï0.05).€Ìò òÌAnalysis€of€agronomic€traitsó óÌ€€€€The€population€means€for€trait€data€based€on€the€combinedÏprogeny€line€means€and€the€progeny€line€means€in€each€of€the€fiveÏenvironments€in€which€SDS€resistance€was€measured€are€shown€inÏTable€1.€The€infection€levels€€as€reflected€by€environment€meanÏdisease€incidence€were€high€(49„64%)€in€all€environments€except€C91Ï(16€%).€However,€the€individual€line€data€in€C91€were€consistentÏwith€environments€in€which€SDS€leaf€symptoms€were€more€severe€(dataÏnot€shown).€For€disease€severity,€environment€means€ranged€from€1.1Ïto€1.7€and€were€broadly€consistent€with€the€DI€environment€means.ÏMean€yield€in€SDS€infested€environments€ranged€from€2410„2810€kg€Ìhaòò„1óó€except€in€C91€(4190€kg€haòò„1óó).€The€higher€yield€in€C91€was€partlyÏa€reflection€of€the€inherent€yield€potential€of€this€particularÏfield€and€partly€of€the€association€of€SDS€leaf€symptoms€withÏreduced€yield€(Gibson€et€al.,€1994).ÌÌò òFrequency€Distribution€Of€SDS€Disease€Incidence.ó óòòóóÌ€€€€€€€€€Mean€DI€òòóóòòóóshowed€an€approximately€normal€(òòPóó€=€0.13),Ïcontinuous€distribution,€although€there€is€evidence€for€a€peakÏbelow€20%€DI€and€a€significant€kurtosis€(„0.65)€reflecting€aÏflattened€distribution€(Fig.€1).€Of€36€lines€with€a€mean€DI€higher€than€Essex€(Fig.€1)€there€were€16€statistically€significantÏsusceptible€transgressive€segregants€(òòPóó€<€0.05).€Ten€lines€had€aÏmean€DI€lower€than€Forrest,€but€none€had€a€significantly€lower€DIÏthan€Forrest.€However,€eight€of€these€lines€were€significantlyÏtransgressive€when€the€LSD€was€determined€from€the€genotype€xÏenvironment€of€the€subset€of€the€10€lines€numerically€superior€toÏForrest€and€excluding€data€from€Forrest.€The€smaller€LSDòò05óó€(4.8)Ïinvolved€in€this€comparison€reflects€the€consistently€low€DI€scoresÏof€these€lines€in€all€environments.€The€eight€transgressive€linesÏmay€be€useful€starting€points€for€developing€soybean€cultivars€withÏstronger€SDS€resistance.Ìò òDNA€Markers€Associated€With€Disease€Incidence.Ì€€€€€ó óTo€detect€loci€conditioning€partial€resistance€to€SDS€weÏtested€associations€between€Fòò5:9óó€genotypic€classes€for€each€DNAÏmarker€and€the€corresponding€mean€SDS€DI€across€five€locations.€AÏsignificant€association€was€declared€if€òòPóó€òò<óó€€0.005€and€if€thatÏmarker€was€linked€to€a€second€marker€that€was€also€associated€withÏthe€trait€with€òòPóó€òò<óó€0.01.€This€minimized€the€detection€of€falseÏassociations€(Landers€and€Botstein,€1989)€and€enabled€QTL€analysis.€Ï€€€€Two€chromosomal€regions€had€significant€effects€on€DI€(TableÏ2).€The€position€of€SDS€resistance€loci€within€each€region€relativeÏto€the€DNA€markers€was€determined€by€QTL€analysis€(Paterson€et€al.,Ï1988)€using€linked€DNA€markers€(Fig.2).€The€first€locus,€identifiedÏby€RAPD€marker€OO05òò250óó,€accounted€for€26%€of€total€variation,€withÏòòPóó€=€0.001€(Table€2).€This€RAPD€marker€mapped€within€6€cM€of€RFLPÏmarker€K455D„1€(òòDraóóI€bands,€Essex€=€5.5€kbp€and€Forrest€=€3.0€kbp).€The€polymorphic€bands€were€generated€with€the€same€restrictionÏenzyme€and€were€of€similar€size€as€K455D„1€that€mapped€to€theÏsoybean€linkage€group€C,€locus€1€(Shoemaker€and€Olson,€1993).ÏTherefore,€the€QTL€was€tentatively€assigned€to€linkage€group€C€ofÏthe€soybean€RFLP€map€(Shoemaker€and€Olson,€1993).€The€K455D„1Ïmarker€was€also€strongly€associated€with€DI€(òòRòòóó2óó€=€0.22,€òòPóó€=€0.001)Ï(Table€3).€The€chromosomal€region€was€flanked€by€distal€andÏproximal€RAPD€markers,€with€neither€significantly€associated€withÏSDS€response,€probably€because€OO05òò450óó€and€OF05òò490óó€mapped€20€and€24€cMÏfrom€OO05òò250óó€and€consequently€were€not€close€to€the€QTL€conditioningÏSDS€response€(Fig.€2).Ì€€€€€€The€second€locus,€identified€by€RAPD€marker€OC01òò650óó,€accountedÏfor€30%€of€the€total€variation€in€DI,€with€òòPóó€=€0.001€(Table€2).€TheÏRAPD€marker€was€mapped€within€14€cM€of€the€RFLP€marker€A071T„1Ï(òòTaqóóI€bands,€Essex€=€null,€Forrest€=€3.0€kbp).€The€Forrest€band€wasÏpolymorphic€with€the€same€restriction€enzyme€and€was€of€similarÏsize€as€the€òòG.maxóó€allele€of€A071„1€that€mapped€to€the€soybeanÏlinkage€group€N,€locus€1€(Shoemaker€and€Olson,€1993).€Therefore,Ïthe€QTL€was€tentatively€assigned€to€linkage€group€N€of€the€soybeanÏRFLP€map€(Shoemaker€and€Olson,€1993).€òòóóThe€RFLP€marker€was€notÏsignificantly€associated€with€DI.€OC01òò650óó€was€linked€to€twoÏadditional€RAPD€markers,€OO04òò1075óó€and€OF04òò1600óó.€€Of€these€only€OF04òò1600óóÏwas€associated€with€DI€(òòRòòóó2óó€=€0.19,€òòPóó€=€0.009).€OF04òò1600óó€mapped€withinÏ14€cM€of€OC01òò650óó.€A071T„1€was€20€cM€and€OO04òò1075óó€28€cM€from€OC01òò650óó€andÏconsequently€were€not€close€to€the€QTL€conditioning€SDS€responseÏ(Fig.€2).Ì€€€€€€òòóóThe€MAPMAKER/QTL€data€suggest€that€together€the€twoÏchromosomal€regions€may€explain€33.6%€of€the€total€variation€in€DI.ÏThe€heritability€of€SDS€DI€in€Essex€x€Forrest€has€been€estimated€atÏ89%€on€a€line€mean€basis€so€that€additional€chromosomal€regionsÏaffecting€SDS€resistance€remain€to€be€discovered€in€this€progenyÏpopulation.€An€analysis€with€seventy€molecular€markers€whichÏencompass€just€1,551€cM€is€unlikely€to€detect€all€SDS€resistanceÏgenes€segregating€in€the€progeny€population.€In€addition,€oneÏhundred€plant€lines€is€a€relatively€small€sample€to€detect€allÏgenes€controlling€SDS€disease€response.€Only€those€partialÏresistance€loci€with€relatively€large€effects€would€be€expected€toÏbe€detected€(Landers€and€Botstein,€1989).Ì€€€€€€The€alleles€at€the€two€mapped€QTL€with€beneficial€effects€onÏDI€were€derived€from€Essex€€at€one€locus€(linkage€group€C)€andÏForrest€at€the€second€locus€(linkage€group€N).€Therefore,Ïresistance€in€Forrest€and€other€resistant€varieties€is€apparentlyÏincomplete€and€can€be€improved,€whereas,€some€SDS€sensitiveÏvarieties,€such€as€Essex,€will€be€useful€sources€of€SDS€resistanceÏalleles€given€marker€assisted€QTL€discovery.€Favorable€alleles€forÏpartial€resistance€to€fungal€pathogens€derived€from€the€susceptibleÏparent€are€commonly€detected€with€molecular€markers€(Bubeck€et€al.,Ï1992;€Wang€et€al.,€1994;€Young€et€al.,€1994).€The€extensiveÏtransgressive€segregation€observed€here€may€result€fromÏrecombination€between€SDS€resistance€alleles€from€both€Forrest€andÏEssex€in€the€progeny€population.ÌÌò òAssociations€Between€Locationsó óÌ€€€€€SDS€infestation€in€the€five€locations€was€consistent€andÏmoderate€to€severe€except€in€C91€where€infestation€was€moderateÏ(Table€1).€The€locus€on€linkage€group€N,€identified€by€OC01òò650óó,€wasÏassociated€with€DI€(òòPóó€<€0.005)€in€four€of€the€five€locations€testedÏ(Table€3).€In€the€fifth€location,€R93,€OC01òò650óó€was€weakly€associatedÏwith€DI€(òòPóó€=€0.008).€The€locus€on€linkage€group€C,€identified€byÏOO05òò250óó,€was€associated€with€DI€(òòPóó€<€0.005)€in€four€of€the€fiveÏlocations€tested€(Table€3).€In€the€fifth€location,€C91€where€DI€wasÏlowest,€OO05òò250óó€was€weakly€associated€with€DI€(òòPóó€=€0.02)€whereas€theÏlinked€marker€K455D„1€was€strongly€associated€with€DI€in€both€C91Ïand€R93€(òòPóó€<€0.0007)€but€was€not€associated€with€DI€in€the€otherÏthree€environments€(òòPóó€>€0.005).€Using€both€OO05òò250óó€and€K455D„1,€theÏpresence€of€the€QTL€could€be€detected€in€all€locations€tested.Ì€€€€€€€The€stability€of€resistance€conferred€by€the€two€chromosomalÏregions€was€clear.€No€evidence€for€consistent€location„specificÏresistance€was€apparent€in€this€genetic€material,€so€no€evidenceÏfor€either€pathologically€distinct€strains€of€òòF.solanióó€in€theÏlocations€tested€or€race„specific€cultivar€resistance€genes€wereÏapparent€from€this€study.€€In€addition€the€Essex€x€Forrest€progenyÏbehave€as€expected€in€SDS€infected€fields€in€Arkansas€(C.€Sneller,Ïpersonal€communication,€University€of€Arkansas,€Fayetteville,€AR,Ï1994).€Therefore,€these€markers€will€be€useful€for€selecting€stableÏSDS€resistant€genotypes€throughout€the€SDS€infested€soybeanÏproduction€area.€The€environmental€stability€of€both€of€these€lociÏcontrasts€with€many€loci€conferring€partial€resistance€to€€fungal€pathogens€of€corn€and€rice€(Bubeck€et€al.€1992;€Wang€et€al€1994).€Ìò òó óÌò òDNA€Markers€Associated€With€Disease€Severityó ó€Ì€€€€€€As€judged€by€the€ANOVA,€the€chromosomal€region€identified€byÏmarker€OC01òò650óó€was€associated€with€DS€(òòRòòóó2óó€=€0.28,€òòPóó€=€0.001).€InÏcontrast€the€chromosomal€region€identified€by€OO05òò250óó€did€not€qualifyÏas€significantly€associated€with€DS€(òòRòòóó2óó€=€0.18,€òòPóó€=€0.007).€NeitherÏchromosomal€region€was€significantly€associated€(LOD€<€2.0)€withÏmean€DS€by€Mapmaker/QTL.€However,€both€QTL€conditioning€SDS€DI€mayÏalso€condition€DS€since€the€greater€significance€of€theÏassociations€with€DI€may€reflect€the€higher€heritability€and€lowerÏenvironmental€interaction€of€DI€compared€to€DS€(Matthews€et€al.,Ï1991;€Gibson€et€al.,€1994;€Njiti€et€al.,€1995).€In€ANOVA,€the€Ïstability€of€marker€associations€between€locations€with€DS€wasÏsimilar€to€those€with€DI,€therefore€DI€was€a€useful€indicator€ofÏeffects€of€these€two€loci€on€DS.€€None€of€the€70€markers€testedÏidentified€loci€strongly€associated€with€DS€that€were€notÏassociated€with€DI.€Therefore,€DI€was€a€useful€indicator€of€theÏeffects€of€all€loci€tested€on€DS.Ìò òÌInteractions€Between€Maturity€Date€And€SDS€Response.€ó óÌ€€€€€SDS€leaf€symptoms€often€do€not€develop€until€several€daysÏafter€flowering€and€in€general€late€maturing€lines€tend€to€be€moreÏresistant€than€early€lines€(Gibson€et€al.,€1994).€Essex€and€ForrestÏare€very€similar€in€maturity€date€and€the€progeny€population€alsoÏshows€a€restricted€variability€in€maturity€date€(Table€1).€There€is€about€10€days€variation€in€date€of€the€R6€stage€(R6€date),€theÏcritical€point€in€SDS€rating.€We€investigated€the€location€of€QTLÏconditioning€the€mean€R6€date€averaged€across€five€locations.€TwoÏunlinked€markers,€OF20òò1025óó€and€OC01òò570óó,€were€strongly€associated€(òòPóó€<Ï0.002)€with€mean€R6€date,€each€accounting€for€27€%€of€theÏvariability.€However,€as€measured€by€Mapmaker/QTL,€neitherÏchromosomal€region€described€above€contained€a€significant€QTL€forÏmean€R6€date€in€SDS€infested€fields€(LOD€<€2).€However,€OF20òò1025óó€andÏOC01òò570óó€were€both€significantly€associated€with€the€R6€date€(òòPóó€<Ï0.005)€in€3€of€the€5€locations€(C91,€P91€and€C92,€data€not€shown).ÏNeither€was€associated€with€SDS€response€nor€linked€to€markersÏencompassing€QTL€conditioning€SDS€response.€Therefore€QTLÏconditioning€the€R6€maturity€date€and€SDS€DI€did€not€correspond,Ïwhich€contrasts€with€the€correspondence€between€QTL€for€partialÏresistance€to€gray€leaf€spot€and€maturity€date€in€corn€(Bubeck€etÏal.,€1992)ÌÌò òDNA€Markers€Associated€with€Yield.ó óÌ€€€€€In€ANOVA,€the€chromosomal€region€identified€by€OC01òò650óó€onÏlinkage€group€N€was€significantly€associated€with€mean€yield€in€SDSÏinfested€test€sites,€accounting€for€27€%€of€variation€(òòPóó€=€0.002,ÏTable€4).€The€association€was€also€significant€in€three€of€fiveÏlocations€(òòPóó€<€0.0008,€Table€4).€Therefore€this€chromosomal€regionÏmay€condition€both€SDS€DI€and€yield,€as€a€result€of€variation€inÏDI,€in€SDS€infested€fields.€The€chromosomal€region€identified€byÏOO05òò250óó€on€linkage€group€C€was€not€significantly€associated€with€mean€yield€(òòPóó€=€0.12)€nor€yield€in€single€locations€(òòPóó€>€0.01).€However,Ïas€measured€by€Mapmaker/QTL,€neither€chromosomal€region€describedÏabove€contained€a€significant€QTL€for€mean€yield€in€SDS€infestedÏfields€(LOD€<€2).€Ì€€€€Significant€but€weak€correlation€coefficients€(r€<€0.6)€betweenÏyield€and€DI€or€DS€have€been€reported€(Gibson€et€al.,€1994).ÏTherefore,€some€loci€which€reduce€the€development€of€leaf€SDSÏsymptoms€(DI€and€DS)€may€not€substantially€improve€yield€potential.ÏSuch€loci€might€reduce€the€symptoms€of€infection€(leaf€chlorosis)Ïwithout€alleviating€the€causes€of€yield€reduction€such€as€root€rot,Ïxylem€plugging€and€flower€and€pod€abortion.Ì€€€€€€The€effects€of€individual€partial€resistance€loci€on€yield€inÏmultiple€environments€have€not€previously€been€reported.€We€haveÏobserved€that€one€locus€associated€with€SDS€resistance€confersÏsignificantly€improved€yield€potential,€whilst€a€second€does€not.ÏThat€differences€are€observed€between€the€two€loci€reported,Ïemphasizes€the€power€of€the€molecular€marker€technique€inÏdissecting€the€components€of€polygenic€disease€resistance.Ìò òÌImplications€for€Genetic€Dissection€and€Marker€Assisted€Breedingó ó€€Ï€€€€€The€markers€K455D„1,€OO05òò250óó,€OC01òò650óó€and€OF04òò1600óó€can€be€used€forÏmarker€assisted€selection€for€partial€SDS€resistance.€This€willÏaccelerate€the€breeding€of€SDS€resistant€soybean€lines€and€mayÏreduce€the€need€for€field€tests€of€SDS€response,€which€are€slow€andÏexpensive.€These€loci€are€currently€being€used€to€determine€allelicÏrelationships€among€different€sources€of€resistance,€to€assist€SDS€resistance€gene€introgression€into€susceptible€germplasm,€and€toÏenable€resistance€gene€pyramiding.Ì€€€€€However,€since€the€currently€identified€favorable€alleles€areÏderived€from€Essex€and€Forrest€their€unequivocal€identification€andÏco„selection€will€be€possible€only€in€restricted€instances,€such€asÏwhen€crossing€with€Essex,€Forrest€or€one€of€their€progeny€lines.ÏThese€loci€may€have€several€alleles,€some€of€which€may€conferÏgreater€or€lesser€SDS€resistance€than€the€alleles€carried€by€EssexÏand€Forrest.€Therefore,€defining€allelic€associations€withÏresistance€at€these€loci€in€a€wider€selection€of€soybean€germplasmÏis€important.€Ì€€€€€In€the€future,€through€genetic€dissection,€the€effects€ofÏsingle€resistance€loci€will€be€determined.€Fòò5:11óó€lines€that€wereÏderived€from€Fòò5óó€plants€heterozygous€in€the€region€of€an€SDSÏresistance€QTL€are€heterogeneous€for€the€region,€whilst€individualÏplants€within€a€line€are€largely€homozygous€within€the€region€byÏthe€Fòò5:11óó.€Lines€heterogeneous€for€each€resistance€locus€have€beenÏidentified€and€sub„line€populations€extracted€from€forty€individualÏplants€within€a€line.€With€these€sub„line€populations€the€geneticÏand€physiological€effects€of€the€segregation€of€an€individualÏresistance€locus€will€be€isolated,€the€region€saturated€withÏmarkers€and€the€QTL€localized€further.€These€studies€will€provideÏa€basis€for€map€based€isolation€of€a€gene€conferring€partialÏresistance€to€SDS.ÌÌò ò€€€€€€€€€€€€€€€€€€€€€€€€€ACKNOWLEDGEMENTSó óÌ€€€€We€thank€Dr€M.€E.€Schmidt€for€developing€the€ExF€progenyÏpopulation.€We€thank€Dr.€R.€Shoemaker€for€the€RFLP€probes€and€someÏparent€RFLP€polymorphism€data.€We€thank€Dr.€P.€Keim€for€much€parentÏRFLP€polymorphism€data.€We€thank€Dr.€D.€Webb€for€some€novel€parentÏRFLP€data€and€verification€of€the€RFLP€polymorphism€data.€We€thankÏall€the€workers€on€the€SDS€field€team€at€Southern€IllinoisÏUniversity€at€Carbondale€from€1987€to€present.€Thanks€to€Dr.€V.ÏNjiti€and€Dr.€O.€Myers€Jr.€for€critical€reading€of€the€manuscript.ÏThis€work€was€supported€in€part€by€grants€from€the€Illinois€SoybeanÏProgram€Operating€Board€Nos.€92„18„125„3,€93„19„132„3,€94„20„143„3.Ç€€€€€€€€€€€€€€€€€€€€€€€€€€€ò òREFERENCESÌó 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òFIGURESó óÌÌFig.€1.€Frequency€distribution€of€the€mean€scores€across€five€fieldÏlocations€for€SDS€disease€incidence€among€100€recombinant€inbredÏlines€from€a€cross€between€cultivars€Essex€and€Forrest.€The€mean€DIÏscore€for€each€parent€in€the€same€environments€is€shown.€The€leastÏsignificant€difference€between€parent€and€line€means€was€16.5€(òòPóó€<Ï0.05).ÌÌFig€2.€Locations€of€RFLP€markers,€RAPD€markers€and€two€QTLÏconditioning€SDS€response.€The€QTL€were€putatively€assigned€toÏlinkage€group€C€and€linkage€group€N€on€the€soybean€genetic€map€byÏanchored€RFLP€(Shoemaker€and€Olson€1993).€END€indicates€the€likelyÏposition€of€the€telomere€on€that€linkage€group,€the€disjunct€barÏindicates€the€rest€of€the€linkage€group.€Marker€names€are€given€onÏthe€left€and€marker„QTL€LOD€scores€are€given€on€the€right€of€eachÏlinkage€group.€LOD€scores€at€markers€were€from€single„locusÏanalyses€of€additive€gene€effects€using€MAPMAKER/QTL€1.1.€GeneticÏdistances€(cM)€were€from€the€recombinant€inbred€line€function€ofÏMAPMAKER/EXP€3.0.€A€cM€distance€scale€is€shown€below€group€C.€TheÏestimated€position€of€the€QTL€is€shown€based€on€2€cM€intervalÏmapping€using€MAPMAKER/QTL€1.1.€€Boxes€indicate€the€region€with€2„¼LOD€(100€fold)€likelihood€intervals.€ÇTable€1.€Means€of€disease€incidence€(DI),€disease€severity€(DS),Ïyield€and€days€to€R6€of€100€Fòò5óó€derived€lines€from€the€cross€EssexÏby€Forrest€tested€in€five€locations.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€ÌóóÌEnv.ð'ð€€€€DI€(%)€€€€€€DS€(1„9)€€€Yield€(kg€haòò„1óó)€€€€€€€Days€to€R6Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌV90€€€€€53€(33)ð(ð€€€€1.7€(0.5)€€€€€2440€(310)€€€€€€€€104.4€(1.4)ÌC91€€€€€16€(19)€€€€€1.1€(0.1)€€€€€4190€(430)€€€€€€€€115.8€(2.2)ÌP91€€€€€57€(29)€€€€€1.4€(0.4)€€€€€2590€(330)€€€€€€€€105.5€(1.5)ÌC92€€€€€64€(25)€€€€€1.5€(0.5)€€€€€2420€(410)€€€€€€€€110.1€(1.6)ÌR93€€€€€52€(30)€€€€€1.6€(0.4)€€€€€2410€(360)€€€€€€€€105.5€(1.9)ÌMean€€€€49€(23)€€€€€1.5€(0.3)€€€€€2810€(290)€€€€€€€€108.1€(1.4)Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÏð'ð€V90,€Villa€Ridge€1990;€C91,€Cora€1991;€P91,€Pulaski€1991;€C92,€€Ï€€Cora€1992;€R93,€Ridgway€1993.Ìð(ð€Standard€deviation€among€progeny€line€means€in€each€location€andÏover€locations€is€shown€in€parentheses.€€€€ÇTable€2.€RAPD€markers€showing€significant€associations€with€meanÏSDS€disease€incidence€across€five€locations.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌ€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€QTL€€€€€€€Mean€DI€%€for€RILsÌ€€DNA€€€Linkage€€€€€€€€€€€€€€€€€€€€€percentage€€with€alleles€fromÌMarker€€groupð'ð€€òòRóóòò2óó€€€€€òòPóó>F€€€€LODð(ð€€variationðð€€Essex€€€€€ForrestÌòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌOO05òò250óó€€€€C€€€€€0.26€€€0.001€€€2.78€€€€18.1€€€€27.3òò+óó7.0€€€€61.0òò+óó6.1Ìà..àà` ..` àÌOC01òò650óó€€€€N€€€€€0.30€€€0.001€€€1.80€€€€15.6€€€€82.3òò+óó2.4€€€€38.5òò+óó7.5Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌð'ð€from€Soybase,€based€on€Shoemaker€and€Olson€(1993).Ìð(ð€LOD€is€the€logòò10óó€of€the€odds€ratio€that€supports€evidence€for€€€€Ï€the€presence€of€the€QTL€at€the€locus€from€Mapmaker/QTL€1.1.Ìðð€the€percent€variation€associated€with€the€interval€fromÏMapmaker/QTL€1.1.€ÇTable€3.€Markers€associated€with€DI€in€single€environments€at€eachÏof€five€locations.€€€€€Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€Ïóó€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€Mean€DI€%€for€linesÌ€DNA€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€with€allele€fromÌòòMarker€€€Env.ð'ð€€€€€òòRóóòò2óóóóòò€€€€€€€òòPóó>F€€€€€€€Essex€€€€€€€€€€€€Fóóòòorrest€€€òòóóóó€ÏÌÌOC01òò650óó€€€V90ภ..¸ à€€0.25àh..hà0.003à..à€90.0€òò+óó4.0à .. à€€€€39.6€òò+óó7.9Ìà..à€€€€C91ภ..¸ à€€0.43àh..hà0.0001à..à€56.0€òò+óó7.0€à .. à€€€€14.0€òò+óó4.1Ìà..à€€€€P91ภ..¸ à€€0.23àh..hà0.004à..à€88.3€òò+óó4.6à .. à€€€€45.0€òò+óó7.2Ìà..à€€€€C92ภ..¸ à€€0.27àh..hà0.002à..à€92.9€òò+óó2.7à .. à€€€€51.0€òò+óó6.3Ìà..à€€€€R93ภ..¸ à€€0.20àh..hà0.008à..à€84.0€òò+óó4.1à .. à€€€€43.0€òò+óó7.4ÌÌK455D„1€€V90ภ..¸ à€€0.15àh..hà0.007à..à€42.9€òò+óó6.7à .. à€€€€67.9€òò+óó5.9Ìà..à€€€€C91ภ..¸ à€€0.23àh..hà0.0006à..à€€8.3€òò+óó2.7à .. à€€€€27.7€òò+óó4.5Ìà..à€€€€P91ภ..¸ à€€0.12àh..hà0.02€à..à€49.1€òò+óó5.5à .. à€€€€68.5€òò+óó1.2Ìà..à€€€€C92ภ..¸ à€€0.08àh..hà0.05àÀ..Àà€€€€€€60.5€òò+óó4.9à .. à€€€€73.4€òò+óó4.2Ìà..à€€€€R93ภ..¸ à€€0.22àh..hà0.0007à..à€40.8€òò+óó6.4à .. à€€€€70.3€òò+óó5.0ÌÌOO05òò250óó€€€V90ภ..¸ à€€0.22àh..hà0.003à..à€27.8€òò+óó9.5à .. à€€€€67.1€òò+óó7.9Ìà..à€€€€C91ภ..¸ à€€0.14àh..hà0.02€€€€à..à€€9.4€òò+óó4.8à .. à€€€€29.1€òò+óó5.7Ìà..à€€€€P91ภ..¸ à€€0.25àh..hà0.001à..à€33.1€òò+óó7.2à .. à€€€€69.5€òò+óó7.1Ìà..à€€€€C92ภ..¸ à€€0.24àh..hà0.002à..à€39.5€òò+óó8.0à .. à€€€€72.1€òò+óó5.7Ìà..à€€€€R93ภ..¸ à€€0.29àh..hà0.0005à..à€26.4€òò+óó8.8à .. à€€€€67.1€òò+óó6.4Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌð'ð€V90,€Villa€Ridge€1990;€C91,€Cora€1991;€P91,€Pulaski€1991;€C92,€€Ï€€Cora€1992;€R93,€Ridgway€1993.ÇÒ X ÒTable€4.€A€Marker€associated€with€yield€in€SDS€infested€fields.ÌÓ  ÓÓ  Óòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌ€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€Mean€yield€(kg€haòò„1óó)Ì€DNA€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€for€lines€with€alleles€fromÌòòMarkeróóòòóóòòóóòò€€€Env.ð'ð€€€€€òòRóóòò2óóóóòò€€€€€€òòPóó>F€€€€€€€€Essex€€€€€€Forrest€€€€€€€€€€€óó€€€€Ï€€ÌOC01òò650óó€€€V90ภ..¸ à€€0.31àh..hà0.0008à..à€2547òò+óó128àÈ..Èà€€€3050òò+óó€60Ìà..à€€€€C91ภ..¸ à€€0.05€àh..hà0.2àÀ..Àà€à..à€4891òò+óó222àÈ..Èà€€€5248òò+óó128Ìà..à€€€€P91ภ..¸ à€€0.10àh..hà0.07àÀ..Ààà..à€2943òò+óó168àÈ..Èà€€€3286òò+óó€87Ìà..à€€€€C92ภ..¸ à€€0.34àh..hà0.0004à..à€2291òò+óó114àÈ..Èà€€€3211òò+óó114Ìà..à€€€€R93ภ..¸ à€€0.31àh..hà0.0007à..à€2399òò+óó141àÈ..Èà€€€3144òò+óó€94Ìà..à€€€Meanภ..¸ à€€0.27àh..hà0.002à..à€3016òò+óó155àÈ..Èà€€€3588òò+óó101Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌð'ð€V90,€Villa€Ridge€1990;€C91,€Cora€1991;€P91,€Pulaski€1991;Ì€€C92,€Cora€1992;€R93,€Ridgway€1993.Çò òó óÓÓÌÌÌÌÌMarch€13,€1995ÌÌDr.€Perry€B.€Cregan,ÌUSDA„ARS,ÌSoybean€and€Alfalfa€Research€Lab.,ÌBldg.€011,€HH„19,ÌBARC„West,ÌBeltsville,€ÌMD20705„2350.ÌÌÌDear€Perry,ÌÌ€€€€€€€€please€find€enclosed€four€copies€of€a€manuscript€for€CropÏSciences€on:€Ìò ò€€€Genetic€Mapping€of€Loci€underlying€Soybean€Sudden€Death€SyndromeÌ€€€€€€€€€€€€€(SDS)€Resistance€and€Tolerance.Ìó óby€€€€€€€N.€Hnetkovsky,€S.€Chang,€T.W.€Doubler,Ì€€€€€€€€€€€P.T.€Gibson€and€D.A.€Lightfootòò*óó.ÌÌ€€€€€€€As€part€of€the€"C7"€experiment€please€send€out€for€review.€Dr.ÏShoemaker€and€Dr€Keim€would€be€appropriate.ÌÌ€€€€€€I€enclose€an€extra€copy€for€consideration€by€the€SoybeanÏGenetics€Committee.€Please€consider€whether€we€are€correct€to€use€SDSòòRóóÏfor€our€QTL€or€should€we€use€òòrfsóó€even€though€the€association€betweenÏSDS€and€òòFusarium€solanióó€is€not€absolute.€A€compromise€might€be€òòrsdóó€forÏresistance€to€sudden€death!ÌÌSincerely,ÌÌÌDavid€A.€LightfootÌAssistant€ProfessorÌÌÌPS.€I€just€received€your€preprint.€Many€Thanks.€Unfortunately€theÏfigures€and€table€were€missing,€could€you€send€them€on?ÇÌÌÌÌÌJune€9,€1995ÌÌDr.€Perry€B.€Cregan,ÌUSDA„ARS,ÌSoybean€and€Alfalfa€Research€Lab.,ÌBldg.€011,€HH„19,ÌBARC„West,ÌBeltsville,€ÌMD20705„2350.ÌÌÌDear€Perry,ÌÌ€€€€€€€€please€find€enclosed€four€revised€copies€of€a€manuscript€forÏCrop€Sciences€as€part€of€the€"C7"€experiment€on:€Ìò ò€€€Genetic€Mapping€of€Loci€underlying€Soybean€Sudden€Death€SyndromeÌ€€€€€€€€€€€€€(SDS)€Resistance€and€Tolerance.Ìó óby€€€€€€€N.€Hnetkovsky,€S.€Chang,€T.W.€Doubler,Ì€€€€€€€€€€€P.T.€Gibson€and€D.A.€Lightfootòò*óó.ÌÌI€appreciated€the€rigorous€but€fair€reviews€and€have€answered€eachÏpoint€and€added€a€few€alterations€of€my€own€in€the€spirit€of€theÏreviewers€comments.€It€was€clearly€in€need€of€revision€partly€beingÏour€first€in€the€subject€area€and€my€first€in€Crop€Science.€TheÏsuggestions€have€been€very€helpful€in€developing€a€standardized€styleÏof€data€presentation.€I€enclosed€the€marked€up€copies€of€the€originalÏand€a€letter€referring€to€your€letter€point€by€point.€€€€€€€Ì€€€€€€€Many€thanks€for€the€material€on€the€assignment€of€geneticÏsymbols.€This€material€has€enabled€us€to€design€appropriateÏexperiments€for€future€development€of€appropriate€gene€symbols.ÌÌÌSincerely,ÌÌÌDavid€A.€LightfootÌAssistant€ProfessorÌÌÌPS.€Polymorphism€with€minisatellites€between€E€and€F€is€about€15/31€soÏfar.ÇNEW€PAGE/POINT€#/OLD€PAGEÌÌReviewer€1ÌÌPage€8€line€18€to€page€9€line€5€#1€Page€7,€lines€16€to€page€8€line€4.ÏThere€were€multiple€replications€per€location€and€multiple€locations.ÏI€hope€the€rewording€of€the€Materials€and€Methods€clarifies€this.ÌÌDeleted€#2,€page€11€lines€8„14.€Clustering€was€a€trend€rather€than€aÏstatistically€significant€phenomenon.€Since€this€is€not€theÏappropriate€data€set€with€which€to€address€this€point€I€will€retractÏit.ÌÌThroughout€#3a.€I€choose€polygenic€to€describe€the€multiple€lociÏaffecting€SDS€response.€However€in€cross€between€a€pair€of€cultivarsÏSDS€response€is€conditioned€only€by€the€segregating€loci€an€so€may€beÏoligogenic€or€even€monogenic.€I€tried€to€be€more€specific€in€usage€andÏused€polygenic€where€no€contrary€evidence€existed.ÌÌDeleted€#3b.€page€1€line€26.€Yes€alleles.ÌÌPage€9€line€10€#3c.€page€8,€line€3.€Defined€and€referenced€at€theÏparagraphs€start.ÌÌ10€line€11€page€11€line€2€#3d.€page€9,€lines€2€and€15.€DoneÏadequately?ÌÌpage€15€line€19€#3e.€page€12,€should€have€been€Fig.2€not€Fig.3.ÌÌpage€15€line€16€#3f.€page€12€line€12.€Modified€to€clarify.ÌÌpage€20€line€18€#3g.€page€15€line€2.€Table€3€not€Table€6.ÌÌpage€21€line€16€#3h.€page€15€line€18.€Specificity€would€be€enhanced€byÏappropriate€genetic€symbols€for€such€QTL,€could€we€arbitrarily€assignÏsymbols?ÌÌpage€24„#3i.€Unfortunately€many€key€references€are€still€onlyÏabstracts.€However€the€list€has€been€shortened€from€59„45€references.ÏNormal€for€a€paper€of€this€wide€scope€(see€Bubeck€et€al.,€1992;€44Ïrefs).ÌÌPage€32„36€#3j.€Table€headings€were€modified€as€suggested,€but€seemÏtypical€of€the€genre.ÌÌDeleted€#3k.€Page€1,€line€22.€We€describe€the€loci€as€resistance€lociÏbut€on€detailed€analysis€we€discovered€the€beneficial€locus€from€EssexÏdid€not€improve€yield€in€any€environment.€Our€measure€of€leaf€symptomsÏis€an€indirect€measure€of€the€disease,€the€fungus€is€restricted€to€theÏtaproot.€Therefore€we€expect€some€"resistance"€loci€do€not€restrictÏfungal€colonization€but€simply€allow€the€beans€to€avoid€leaf€chlorosisÏbut€not€the€yield€loss.€I€was€incorrect€to€use€tolerance,€however.̇Reviewer€2ÌÌPage€3€line€2€page€6€line€24€#1€There€are€as€yet€no€clear€statementsÏregarding€the€stability€of€SDS€resistance,€or€existence€of€racesÏalthough€cross€regional€studies€are€in€hand.€In€lieu€of€these€resultsÏI€modified€the€abstract€(Page€3€line€2)€and€reorganized€theÏintroduction€(page€5€line€1„6€moved€to€page€6€line€24)€on€this€point.ÌÌpage€12€line€11€#2€Page€10€line€16.€QTL€analyses€on€RIL€mapping€dataÏmay€be€undertaken€by€converting€the€data€file€to€F2€backcross.ÌÌDeleted€#3€Page€11€lines€8„14.€Clustering€was€a€trend€rather€than€aÏstatistically€significant€phenomenon.€Since€this€is€not€theÏappropriate€data€set€with€which€to€address€this€point€I€will€retractÏit.ÌÌpage€16€line€23€to€page€17€line€9€#4€Page€12,€line€5€Our€criteria€wereÏmore€stringent€than€reported.€Stringent€criteria€for€P€AND€linkage€ofÏseveral€marker€led€us€not€to€report€on€3€additional€putative€loci,Ïonly€the€two€best.€We€felt€clarity€was€improved€also.€(lines€7„11Ïmoved€to€end€add€seventy€marker€story)€The€small€population€size€andÏlow€marker€density€undoubtedly€led€some€resistance€QTL€to€be€missed.€ÌÌpage€16€line€5„7€#5€Page€13€lines€7,8.€Good€point.ÌÌThroughout€#6€Page€9€line€14.€Good€Point.ÌÌThroughout€#7.€Page€2€line€2.€Done.ÌÌpage€2€line€22#8.€Good€point.ÌÌ#9.€Page€4€line€6„9€#9a€Page€4€line€3„7,Ì€€€€Page€4€line€20„22€#9b€p€4€line€19„20,Ì€€€€page€14€line€21„24€#9c€page€11€line€21„24,€Ì€€€€page€18€line€26€#9d€page€14€line€13„16.€Clarified€but€less€€€Ïconcise.ÌÌPage€6€line€2„8€#10€Page€6,€lines€7„19.€I€strongly€disagree€that€cystÏnematode€resistance€has€been€proven€to€be€a€race„insensitive€partialÏresistance€hence€it€was€excluded€from€the€original€paragraphÏ(Concibido€et€al.€1994,€Kreike€et€al€1994).€I€have€included€aÏparagraph€(with€references)€and€altered€the€wording€in€another€toÏinclude€the€dilemma€that€SCN€resistance€seems€to€represent€ie.€partialÏresistance€that€is€HIGHLY€race€specific.€There€was€one€good€referenceÏto€quantitative€inheritance€of€field€resistance€to€a€fugal€pathogen€inÏcorn€(Bubeck€et€al.€1994).€There€were€2€good€references€toÏquantitative€resistance€to€bacterial€pathogens€I€overlooked€and€IÏadded€one€more€recent€example.€However€all€this€contradicts€theÏefforts€to€reduce€the€reference€list!ÌÌPage€14€line€23„26€#11.€Page€11€lines€20„25.€We€added€the€LSD€here€andÏto€the€figure.€The€reasons€for€lack€of€significance€of€the€positive€transgressive€segregants€is€discussed€here€and€on€page€13€line€15€asÏwas.€Also€discussed€on€page€17€line€21.ÌÌpage€11€line€23€#12.€Added€a€line€to€end€of€the€RAPD€section€inÏMaterials€and€Methods€(Page€10€line€4).€RAPD€markers€are€reproducibleÏif€you€take€care€to€reproduce€the€conditions€and€reagents€exactly.€WeÏhave€cloned€the€important€RAPD€bands€and€are€in€the€process€of€mappingÏas€RFLPs€or€ASAPs.€In€addition€their€positions€relative€to€publiclyÏavailable€RFLPs,and€SSRs€can€be€inferred€from€the€data€presented!ÌÌÌpage€13€line€22€#13.€Added€to€page€11€line€14.€ÌÌpage€32€#14.€Decided€against€combining€tables€1€and€2€for€clarity.ÌÌDeleted€#15.€Crosses€Pyramid€x€Douglas€and€Williams€x€Williams€82.ÌÌReviewer€3ÌÌPage€4€line€26€#3,€Page€4,€line€26.€We€altered€the€wording€somewhat.ÌÌPage€6€line€2€#4,€Page€6,€lines€7„19.€I€strongly€disagree€that€SCNÏresistance€has€been€proven€to€be€a€race„insensitive€partial€resistanceÏhence€it€was€excluded€from€the€original€paragraph.€I€have€included€aÏparagraph€(with€references)€and€altered€the€wording€in€another€toÏinclude€the€dilemma€that€SCN€resistance€seems€to€represent€ie.€partialÏresistance€that€is€HIGHLY€race€specific€(except€in€one€case).ÌÌPage€8€line€18€to€page€9€line€5#5€See€reviewer€1€#1.ÌÌÌPage€9€line€10#6€Page€8,€lines€3„5.€I€have€included€a€description€ofÏthe€DS€scale€used.ÌÌPage€10€line€9€#7€Page€8,€lines€21„24.€I€have€reorganized€for€clarityÏas€suggested.ÌÌPage€14€€line€1„15€page€32€#8€a,€page€11€lines€8„14.€We€added€aÏparagraph€and€a€table.€Yes€the€lattice€design€was€more€efficient€thanÏan€RCB.€ÌÌDeleted€#9€a.€page€11€lines€8„14.€Clustering€was€a€trend€rather€than€aÏstatistically€significant€phenomenon.€Since€this€is€not€theÏappropriate€data€set€with€which€to€address€this€point€I€will€retractÏit.ÌÌPage€13€line€9€#9b.€page€11€line€10.€Wording€is€altered.ÌÌPage€13€line€13€#9c.€page€11€line€14.€Appended.€Since€the€clusteringÏstatements€are€withdrawn€showing€the€linkage€groups€would€beÏredundant.€There€were€only€eight€groups€not€ten€when€applying€theÏcriteria€set€forth€in€materials€and€methods.̇Throughout€#10.€Corrected€ròò2óó€to€Ròò2óó€throughout.ÌÌPage€14€line€21€#11.€Page€11€lines€20„25.€We€added€the€LSD€here€and€toÏthe€figure.€The€lack€of€significance€of€the€positive€transgressiveÏsegregants€is€discussed€here€and€on€page€13€line€15€as€was.€WeÏcalculated€LSD€to€indicate€which€transgressive€segregants€wereÏstatistically€significant.€In€fact€the€parents€were€extensivelyÏreplicated.€It€is€likely€there€are€additional€transgressive€segregantsÏthat€can€be€revealed€by€a€more€sophisticated€statistical€analysis,Ïthis€will€be€part€of€a€paper€on€the€field€data€(without€DNA€markers).ÌÌPage€15€line€19€#12.€p12€line€15,.€Corrected€to€Fig€2ÌÌPage16€line€5,€throughout€#13.€p13€lines4„5.€RAPD€markers€were€linked,Ïidentified€but€did€not€define€chromosomal€region€conditioning€SDSÏresponse.ÌÌPage€20€line€20€#14.€p€15€line€2.€Table€3€correct€(not€6).ÌÌPage€21€line€13€#15.€p15€line€13„5.€Added€a€sentence€to€clarify.ÌÌPage€21€line€19€#16.€p15€line€19.€DeletedÌÌPage€22€line€1,€#17.€p16€line1„3.€We€were€making€the€point€that€weÏonly€know€the€relative€benefits€of€two€alleles€at€each€locus.ÏTherefore,€we€only€know€which€allele€to€select€in€crosses€with€EssexÏand€Forrest€or€their€progeny€lines.€I€hope€this€is€clarified€in€theÏtext.€It€is€intended€as€a€cautionary€statement€to€those€interested€inÏmarker€assisted€selection€with€the€markers!€We€need€unequivocalÏminisatellite€identification€of€these€alleles!€ÌÌPage€22€line€11€#18.€p16€line€9.€I€misspelled€heterogenic€(EnglishÏstyle).€The€definition€of€it€is€"Consisting€of€or€involving€dissimilarÏparts;€not€homogeneous".€Adjective.€First€used€to€describe€aÏpopulation€by€Lewis€1947,€Competition€and€dominance€of€incompatibilityÏalleles€in€pollen.€Heredity€1:85„95.€However€since€more€US€breedersÏwill€understand€heterogeneous€I€have€converted€the€text€throughout.ÌÌPage€32€„36€#19.€Tables:€a,€DoneÌ€€€€€€€€€€€€b,€QTL€mapmaker€estimation€of€variation€explained,€this€is€Ï€€€€€€€€€€€€€€€noted€in€the€text.Ì€€€€€€€€€€€€c,€RevisedÌPage€31,€37,€38€#20.€Figure:€RevisedÌ€€€€€€€€€€€€a,€END€refers€to€the€telomere€or€end€of€the€existing€€€€€€€Ï€€€€€€€€€€€€€€€linkage€group.€Revised€Legend.Ì€€€€€€€€€€€€b,€Drawings€are€to€scale,€were€in€error.Ì€€€€€€€€€€€€c,€these€are€only€fragments€of€the€linkage€group,€our€70€€€Ï€€€€€€€€€€€€€€€€marker€map€was€fragmentary.Ì€€€€€€€€€€€€d,€Revised€as€suggested€A71T„1€is€now€only€included,€€€€€€€Ï€€€€€€€€€€€€€€€recent€data€suggests€the€linkage€of€A71c€(A71T„3)€was€a€Ï€€€€€€€€€€€€€€€transitive€mapping€error.Ìpage€24„#21.€Unfortunately€many€key€references€are€still€onlyÏabstracts.€However€the€list€has€been€shortened€from€59„45€references.ÏNormal€for€a€paper€of€this€wide€scope€(see€Bubeck€et€al.,€1992;€44Ïrefs).ÌÌYour€commentsÌÌ#1€Adopted€throughoutÌÌ#2€Title€revisedÌÌ#3€Abbreviations€addedÌÌ#4€Abstract€modified,€thats€the€molecular€biology€training€showingÏagain,€esoteric€research!ÌÌ#5€We€have€extensively€reviewed€the€refernces,€I€can€find€no€moreÏerrors.ÌÇRemoved€from€Abstract.Ì€Detection€of€a€beneficial€allele€in€'Essex',€the€susceptible€parent,€Ïdemonstrates€the€potential€of€marker€analysis€in€gene€discovery€andÏintrogression.€We€identified€15€Fòò5óó€derived€lines€containing€beneficialÏalleles€at€both€loci,€some€of€€These€lines€may€be€useful€in€soybeanÏvariety€development.ÇOF04òò1600óó€€€V90ภ..¸ à€€0.19àh..hà0.009à..à€41.5òò+óó1.4€€€€46.2òò+óó1.0€àÐ ..Ð à€€€€€€€€€€€Ïà..à€€€€C91ภ..¸ à€€0.04àh..hà0.23àÀ..Àà€€€€€€75.5òò+óó2.5€€€€78.8òò+óó1.4€Ìà..à€€€€P91ภ..¸ à€€0.06àh..hà0.17àÀ..Àà€€€€€€47.4òò+óó1.7€€€€50.1òò+óó1.1€Ìà..à€€€€C92ภ..¸ à€€0.15àh..hà0.02àÀ..Àà€€€€€€41.7òò+óó2.2€€€€48.4òò+óó1.7€Ìà..à€€€€R93ภ..¸ à€€0.37àh..hà0.0001à..à€39.2òò+óó1.8àÈ..Èà€€€48.4òò+óó1.2Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌ*€V90,€Villa€Ridge€1990;€C91,€Cora€1991;€P91,€Pulaski€1991;Ì€€C92,€Cora€1992;€R93,€Ridgway€1993.ÇTable€2.€RAPD€markers€and€linkage€groups€showing€significantÏassociations€with€mean€SDS€disease€incidence€across€five€locations.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌ€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€QTL€€€€€€€Mean€DI€for€linesÌ€€DNA€€€Linkage€€€€€€€€€€€€€€€€€€€€€€€percentage€€with€alleles€fromÌ€Marker€€groupð'ð€€€òòRóóòò2óó€€€€€òòPóó>F€€€€LODð(ð€€variationðð€€Essex€€€ForrestÌòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌOO05òò250óó€€€€C€€€€€0.26€€€0.001€€€2.78€€€€18.1€€€€€€27.3òò+óó€€€€61.0òò+óóÌà..àà` ..` àÌOC01òò650óó€€€€N€€€€€0.30€€€0.001€€€1.80€€€€15.6€€€€€€82.3òò+óó€€€€38.5òò+óóÌòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌð'ð€from€Soybase,€based€on€Shoemaker€and€Olson€(1993).Ìð(ð€LOD€is€the€logòò10óó€of€the€odds€ratio€that€supports€evidence€for€€€€€Ïthe€presence€of€the€QTL€at€the€locus€from€Mapmaker/QTL€1.1.Ìðð€the€percent€variation€associated€with€the€interval€from€Mapmaker/QTLÏ1.1.ÇTable€1:€RAPD€markers€and€Linkage€groups€showing€significantÏassociations€with€SDS€response.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌ€€€€€€€Linkage€€€€€€€€€€€€€€€€€€€€€€€Percentage€€€Allelic€means€Probe€€Ïgroup€€€€Ròò2óó€€€€€P>f€€€€LODòò*óó€€€variation€€€€€E€€€€€€€€FÌòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌa,€Disease€IncidenceÌOPO05e€€C€€€€€€€0.26€€€0.001€€€2.78€€€€18.1€€€€€€€27.3€€€€61.0Ìà..àà` ..` àÌOC01òò650óó€€N€€€€€€€0.30€€€0.001€€€1.80€€€€15.6€€€€€€€82.3€€€€38.5ÌÌÌb,€Disease€SeverityÌOPO05e€€C€€€€€€€0.18€€€0.007€€€€ND€€€€€ND€€€€€€€€€1.37€€€1.72€€€ÌÌOC01òò650óó€€N€€€€€€€0.28€€€0.001€€€€ND€€€€€ND€€€€€€€€€2.02€€€1.47€€€€€ÌÌÌc,€YieldÌOPO05e€€€€€€€€€0.07€€€€0.12€€€€€ND€€€€€ND€€€€€€€€€53.5€€€50.3€€€à(#..(#àà€%..€%àÌOC01òò650óó€€N€€€€€€0.27€€€€0.002€€€€ND€€€€€ND€€€€€€€€€44.9€€€53.4€Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€ÌóóLOD€is€the€log€10€of€the€odds€ratio€that€supports€evidence€for€theÏprescence€of€the€QTL€at€the€locus.€ND€indicates€no€QTL€was€detected€inÏthis€chromosomal€segment€(LOD€<€2.0).ÌTable€5:€Interactions€between€loci€conditioning€SDS€response.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€óó€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€Ì€€€€€€€€€€€€€€€€OC01òò650óóÌ€€€€€€€€€€€€€E€€€€€€€€€€F*Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌ€€€€€€€€E*€€81.3€€€€€€€15.0ÌSOP05eÌ€€€€€€€€€F€€82.58€€€€€€55.46Ó&€..XÐd&ÓÇTable€1:€RAPD€markers€and€Linkage€groups€showing€significantÏà0..àà0` ..` àassociations€with€SDS€response.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌ€€€€€€€Linkage€€€€€€€€€€€€€€€€€€€€€€€Percentage€€€Allelic€meansÌProbe€€group€€€€€Ròò2óó€€€€€P>f€€€€LODòò*óó€€€variation€€€€€€E€€€€€€€FÌòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌOO05òò250óó€€€€€C€€€€0.26€€€0.001€€€2.81€€€€18.1€€€€€€€27.3€€€€61.0Ìà..àà` ..` àÌOC01òò650óó€€€€€N€€€€0.30€€€0.001€€€1.80€€€€15.6€€€€€€€82.3€€€€38.5ÌÌSPE04h€€€€G€€€€€0.39€€€0.001€€€4.16€€€€25.3€€€€€€€65.2€€€€28.3ÌÌSPG13a€€€€A€€€€€€€0.36€€€0.0001€€3.96€€€€26.5€€€€€€€65.1€€€€14.3€ÌÌSPF02c€€anon2€€€0.14€€€0.003€€€2.12€€€€13.9€€€€€€€40.7€€€€60.4òòÌ€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌLOD€is€the€log€10€of€the€odds€ratio€that€supports€evidence€forÌ€€€€€€€€the€prescence€of€the€QTL€at€the€locus.€ÌÇÔ€–––ÔÔS€/-xÐðA`&UniversScalableSÔÔ€dddÔÔS€0,xÐðA`&UniversScalableSÔTable€1:€RAPD€markers€and€Linkage€groups€showing€significant€Ìassociations€with€SDS€response.Ìòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌ€€€€€€€Linkage€€€€€€€€€€€€€€€€€€€€€€€Percentage€€€Allelic€meansÌProbe€€group€€€€€Ròò2óó€€€€€P>f€€€€LODòò*óó€€€variation€€€€€€E€€€€€€€FÌòò€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌOPO05e€€C€€€€€€€0.26€€€0.001€€€2.78€€€€18.1€€€€€€€27.3€€€€61.0Ìà..àà` ..` àÌOPC01k€€N€€€€€€€0.30€€€0.001€€€1.80€€€€15.6€€€€€€€82.3€€€€38.5ÌÌSPF04h€€G€€€€€€€0.39€€€0.001€€€4.16€€€€25.3€€€€€€€65.2€€€€28.3ÌÌSPG13a€€F€€€€€€€0.36€€€0.0001€€3.96€€€€26.5€€€€€€€65.1€€€€14.3€ÌÌSPF02c€€B€€€€€€€0.14€€€0.003€€€2.12€€€€13.9€€€€€€€40.7€€€€60.4òòÌ€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€óóÌLOD€is€the€log€10€of€the€odds€ratio€that€supports€evidence€forÌ€€€€€€€€the€prescence€of€the€QTL€at€the€locus.€ÌÇÑ  ÑÑ  ÑÑ\R3Ø'\ÑÔ€XXXÔÔO€.<þ6X9`("Courier 10cpiOÔ€€€b,€ÇSignificant€Loci€for€Epistasis€TablesÌÌ€€€€€€€€SOF05e€OPC01k€SF04h€SF02c€SG13aÌSOF05e€ÌOPC01k€ÌSF04hÌSF02cÌSG13aÌÌAny€loci€with€5€beneficial€alleles?€Whats€their€mean€DIÌÌÌÌA117€A847€K455?ÌÌÇSimon€for€the€first€paper€I've€settled€on€the€five€most€stronglyÏassociated€loci€to€analyze€which€include€3€you€just€discovered.€ÌÌStrongly€Significant€Unlinked€Loci€for€SDS€response€(P<0.005).ÌÌSOF05e€(C)ÌOPC01k€(N)ÌSF04h€(?)€ÌSF02c€(F€or€N)ÌSG13a€(?)ÌÌEach€is€linked€to€at€least€1€other€marker€so€Mapmaker€analysis€may€beÏpossible€in€addition€to€ANOVA.ÌÌWe€need€ASAPÌ1.€The€r2€P€and€allelic€means€for€mean€DI.€ANOVA.Ì2.€A€revised€estimate€of€joint€heritability.€ANOVA.Ì2.€A€table€of€r2,€P€and€QTL€LOD€for€DI€in€each€environment.€BOTH.Ì3.€A€comparison€of€r2,€P€and€LOD€for€DI,€DS,€and€Yield€means.€BOTH.ÇÇStrain€€€€€Species€€€€€€€€€€€€€€Host€€€€€€€€SDS€IncitantÌÌX€€€€€€€€€Septoria€?€€€€€€€€€€€€€€Various€€€€€€€€€€€NoÌT8€€€€€€€€Fusarium€solani€type€A€Alfalfa?€€€€€YesÌD1€€€€€€€€Fusarium€solani€type€A€Alfalfa?€€€€€€YesÌ1177€€€€€€Fusarium€solani€type€A€€soybean€€€€€€Yes€€€€€€€€€€€€€€€€€91„¼10„1ÌMont€€IlÌRW1„A€€IlÌVR4€€IlÌST90€€IlÌ269€€ArkÌ17„1€€ArkÌMal1792€€MOÌFSA1€€MSÌÇò òInteraction€between€SDS€response€loció óÌ€€€€€The€interaction€of€the€SDS€response€loci€defined€SOP05e€andÏOPC01k€is€clearly€not€additive.€Epistasis€of€OPC01k€over€OPO05e€isÏapparent€(Table€4).€The€favourable€allele€of€OPC01k€was€present€amongÏthe€resistantprogeny€more€frequently€than€expected€by€chance€(P€<Ï0.01)€emphasizing€the€importance€of€this€allele€in€conditioning€SDSÏresponse.Ìò òó ó€€€€€The€combined€effects€of€pairs€of€loci€is€illustrated€by€Table€2.ÏNote€that€individuals€that€contained€beneficial€alleles€at€SOP05eÏ(group€C)€and€OPC11k€(group€N)€had€SDS€DI€means€equal€to€or€lower€thanÏForrest€the€agronomically€resistant€parent.€Therefore,€just€two€ofÏthese€five€resistance€loci€may€be€sufficient€to€confer€agronomicallyÏadequate€SDS€resistance€on€a€variety.€Combining€all€five€resistanceÏloci€in€a€single€genotype€provides€a€very€high€level€of€resistanceÏ(Table€3).ÌÇ€SDS€resistance€is€associated€with€cyst€nematode€race€3€resistance€inÏadapted€soybean.€Forrest€derives€SCN€race€3€resistance€from€PekingÏwhereas€Essex€is€SCN€susceptible€allowing€the€lines€to€be€assayed€forÏsoybean€cyst€nematode€disease€response.€Assays€in€the€greenhouse€usedÏsoil€infested€with€òòHeterodera€glycineóó€that€tested€as€race€3€fromÏElkville,€Illinois€to€contrast€disease€response€and€molecular€markers.ÏSDS€and€SCN€resistance€loci€were€found€to€be€closely€linked€at€threeÏloci€and€independent€at€two€others.€The€implication€is€that€selectionÏof€SCN€resistance€free€of€linkage€drag€might€increase€SDSÏsusceptibility€in€adapted€soybean€germplasm.ÌÌÌ€€€€€SOYBEAN€SUDDEN€DEATH€SYNDROME€(SDS)€is€causing€increasinglyÏsevere€losses€to€soybean€production€(Mulrooney€et€al€1988,€Gibson€etÏal€1993,€Yang€and€Rizvi€1994).€To€date,€host€plant€resistance€has€beenÏthe€most€effective€control€measure€(Brown€1987)€but€it€is€difficult€toÏdetermine,€incomplete€(partial),€oligogenic€in€nature€and€geneticallyÏcomplex.€Classical€genetic€studies€indicate€that€resistance€isÏconditioned€by€more€than€five€major€genes€(Matthews€et€al.€1991,€NjitiÏet€al.1991).€òòFusarium€solanióó€type€A€has€been€shown€to€be€the€incitantÏof€sudden€death€syndrome€(Rupe€et€al€1989,€Roy€et€al€1989)€and€geneticÏstudies€in€greenhouse€assays€indicate€resistance€to€leaf€symptomÏexpression€may€be€conditioned€by€a€single€gene,€òòrfsóó,€in€one€caseÏ(Stephens€et€al.€1993).€However,€in€the€field€òòrfsóó€does€not€confer€SDSÏresistance,€in€fact€all€SDS€resistant€genotypes€tested€to€date€showÏloss€causing€disease€symptoms€during€conditions€conducive€to€theÏdisease€expression€(Gibson€et€al€1991).€Consequently€useful€fieldÏresistance€to€SDS€is€expected€to€be€oligogenic€in€nature.€€ÌÌ€€€€€€Partial€resistance€to€SDS€is€unlikely€to€represent€the€classicalÏrace„cultivar€specific€resistance€manifest€in€the€hypersensitiveÏresponse€rather€it€will€reflect€the€additive€effect€of€interactionsÏbetween€many€genes€(Njiti€et€al€1991,€Matthews€et€al€1991).€PartialÏresistance€may€be€preferable€to€single€gene€resistance€since€it€mayÏprovide€stable€resistance€to€genetically€variable€field€populations€ofÏòòFusarium€solanióó€type€A.ÌÌ€€€€€€A€strong€association€between€SCN€race€3€resistance€and€partialÏSDS€resistance€has€been€observed€among€over€1000€soybean€varietiesÏtested€in€the€field€(Anders€et€al€1989,€Rupe€et€al€1989b,€Gibson€et€alÏ1991).€In€inheritance€studies€between€SCN€race€3€resistant€andÏsusceptible€cultivars€this€association€has€been€shown€to€be€highlyÏheritable€in€two€crosses€(Matthews€et€al€1991,€Njiti€et€al.€1991)€andÏto€account€for€up€to€50%€phenotypic€variability€in€SDS€resistance€inÏthose€cases.€However,€some€SCN€race€3€resistant€cultivars€are€very€SDSÏsusceptible€and€the€original€sources€of€SCN€race€3€resistance,€Peking,ÏPI88.788€and€PI437.654€are€all€very€SDS€susceptible.€Therefore,€duringÏbreeding€selection€for€SDS€resistance€by€SCN€race€3€resistance€isÏunreliable€and€confers€only€a€fraction€of€the€resistance€whenÏeffective.ÌÌ€€€€€€€Breeding€directly€for€SDS€resistance€is€complicated€by€the€abscence€of€a€satisfactory€quantitative€greenhouse€assay,€therebyÏnecessitating€extensive€field€selections€with€many€lines.€Factors€suchÏas€rainfall,€soil€fertility€(Herschman€et€al.€1990,€Rupe€et€al.€1994),Ïmaturity€date,€date€of€planting,€plant€genotype€and€cyst€nematodeÏinfestation€(Anders€et€al.€1989,€Rupe€et€al€1989b)€all€significantlyÏalter€the€degree€of€disease€progression€in€the€field.€Therefore,Ïreliable€field€evaluations€require€multiple€relplications€in€infestedÏsites€with€accurate€disease€progression€data€collected€at€weeklyÏintervals€(Gibson€et€al€1991).€Consequently,€conventional€breeding€forÏSCN€resistance€is€time€consuming,€difficult€and€expensive.ÌÌ€€€€€€€€Molecular€biotechnology€makes€it€possible€to€identifyÏchromosomal€segments€containing€agriculturally€important€genes,Ïincluding€SDS€and€resistance€genes,€with€DNA€markers.€With€DNA€markersÏwill€be€possible€to€rapidly€identify€SDS€resistant€soybean€lines€basedÏon€genotype€instead€of€phenotype.€This€might€eliminate€or€reduce€theÏneed€for€replicated€field€tests.€Potentially,€lines€containingÏcombinations€of€different€types€of€SDS€resistance€loci€could€beÏselected.€In€addition,€molecular€markers€can€dissect€the€relationshipÏbetween€SDS€and€SCN€resistance€into€pleitrophy,€linkage€and€epistaticÏinteractions.ÌÌ€€€€€€€€€In€studying€disease€resistance€in€plants€the€use€of€DNAÏmarkers€has€focussed€on€genes€involved€in€the€race„cultivar€specificÏhypersensitive€response.€These€include€single€gene€resistance€toÏfungal€pathogens€in€tomato€(Young€et€al.€1888,€Martin€et€al€1991,ÏSarfati€et€al€1989);€lettuce€(Michelmore€et€al.€1991);€rice€(Yu€et€alÏ1991);€thrale€(Bent€et€al€1994)€and€soybean€(Diers€et€al€1991,€PolzinÏet€al€1994).€Similarly,€gene€resistances€for€nematode€pests€have€beenÏidentified€in€€tomato€(Messeguer€et€al€1991);€sugar€beets€(Jung€et€alÏ1990)€wheat€(Rivoal€et€al€1994)€and€potato€(Barone€et€al€1990).€InÏsoybean,€SCN€race€3€resistance€DNA€markers€were€used€to€map€threeÏgenes€(Weiseman€et€al€1992,€Concibido€et€al€1994,€Webb€et€al€1995)Ïrequired€for€a€hypersensitve€type€reaction€(Ende€1964).ÌÌ€€€€€€€€DNA€markers€have€not€been€widely€used€in€studying€partialÏdisease€resistance€based€on€interactions€between€several€genes€inÏplants.€The€few€published€include€partial€resistance€to€fungalÏpathogens€in€mung€bean€(Young€et€al€1994)€and€rice€(Wang€et€al€1994).ÏHowever€breeding€for€partial€resistance€to€fungal€pathogens€isÏrelatively€common€because€of€the€stable€nature€of€the€diseaseÏresistance€(Eddington€et€al€1994,€Gingera€et€al€1994,€Newton€et€alÏ1993,€Kari€et€al€1994,€Miklas€et€al€1992,€Shew€et€al€1989)€and€crossÏprotection€afforded€(Zhang€et€al€1994).€Partial€disease€resistance€isÏalso€involved€in€systemic€acquired€resistance€(Ward€et€al.€1991)€whichÏis€mediated€in€part€by€genes€confering€partial€resistance€to€fungalÏpathogens€(Alexander€et€al€1994,€Cornellissen€et€al€1993,€Sela„¼Burlinger€et€al€1993).ÌÌ€€€€€€€€€In€this€paper€we€demonstrate€the€utility€of€DNA€geneticÏmarkers€for€identifying€chromosomal€segments€contributing€partialÏresistance€to€SDS€and€separating€them€from€loci€conditioning€SCN€race€3€resistance.€The€implications€for€the€use€of€DNA€markers€for€improvedÏselection€of€SDS€resistance€in€soybean€are€considered.ÌÌÌÒ ° ÒBibliographyÌAnders,€M.L.,€Gibson,€P.T.€and€Myers€Jr,€O.€(1989).€Comparisons€¼à0..àof€soybean€cultivar€resistance€to€SDS€and€SCN.€1989€Agron.ÏAbstr.€p72.Ì3.€Andreas,€K.G.€&€Griffiths€E.€(1993).€Components€of€partial€¼à0..àresistance€of€barley€to€òòRhynchosporium€secalisóó:€Use€ofÏseedling€tests€to€prdict€field€resistance.€Annals€of€AppliedÏBiology€123:545Ì4.€Alexander,€D.,€Goodman,€R.M.,€Gut„Rella,€M.,€Glascock,€C.,€¼à0..àWeyman,€K.,€Friedrich,€L.,€Maddox,€D.,€Ahl„Goy,€P.,€Luntz,ÏT.,€Ward,€E.€&€Ryals,€J.€(1993).€Increased€tolerance€to€twoÏoomycete€pathogens€in€transgenic€tobacco€expressingÏpathogenesis„related€protein€1a.€Proc.€Natl.€Acad.€Sci.€USAÏ90:7327„7331.ÌÌBarone,€A.,€E.€Ritter,€U.€Schachtschabel,€T.€Debener,€F.€SalaminiÏand€C.€Gebhardt.€1990.€Localization€by€restriction€fragmentÏlength€polymorphism€mapping€in€potato€of€a€major€dominant€geneÏconferring€resistance€to€the€potato€cyst€nematode€òòGloberiaÏrostocherensisóó.€Mol.€Gen.€Genet.€224:177„182.Ì6.€Bent,€A.F.,€Kunkel,€B.N.,€Dahlbeck,€D.,€Brown,€K.L.,€Schmidt,€¼à0..àR.,€Giraudat,€J.,€Leung,€J.€&€Staskawicz,€B.J.€(1994).€RPS2Ïof€Arabidopsis€thaliana:€A€leucine€rich€repeat€class€ofÏplant€disease€resistance€genes.€Science€265:1856„1859.Ì8.€Brown,€R.H.€(1987).€Control€strategies€in€low€value€crops.€¼à0..àpp179„232.€In€R.H.€Brown€and€B.H.€Kerry€(ed.)€Principles€andÏpractice€of€nematode€control€in€crops.€Academic€Press,ÏOrlando€Fl.Ì13.€Cornelissen,€B.J.C.,€Melchers,€L.S.€(1993).€Strategies€for€¼à0..àcontrol€of€fungal€disease€with€transgenic€plants.€PlantÏPhysiol.€101:709„712.ÌÌòòóó12.Concibido,€V.,€Boutin,€S.,€Denny€R.,€Hautea€R.,€Orf,€J.€Young,Ïà0..àN.D.€(1994).€DNA€marker€analysis€of€loci€underlyingÏresistance€to€soybean€cyst€nematode€òò(Heterodera€glycineóóÏIchinohe).€Crop€Sci.€34:240„246.Ì15.€Diers,€B.W.,€Mansur,€L.,€Imsande,€J.€Shoemaker,€R.€(1992)€¼à0..àMapping€Phytophtora€resistance€loci€in€soybean€withÏrestriction€fragment€length€polymorphism€markers.€Crop€Sci.Ï32:377„383.ÌEddington,€B.R.,€Shanahan,€P.E.€and€Rijkenberg,€F.H.J.€1994.ÏBreeding€for€partial€resistance€in€dry€beans€(òòPhaseolus€vulgarisóóÌto€bean€rust€(òòUromyces€appendiculatusóó).€Anal.€Appl.€Biol.Ï124:341„Ì17.€Endo,€B.Y.€(1964).€Penetration€and€development€of€òòHeterodera€¼à0..àglycinesóó€in€soybean€roots€and€related€anatomical€changes.ÏPhytopathology€54:79„88.Ì20.€Fehr,€W.R.€Caviness,€C.E.,€Burmood,€D.T.,€Pennington,€J.S.€à0..à(1971).€Stages€of€development€descriptions€for€soybeans.ÏCrop€Sci€11:929„931.Ì23.€Gibson,€P.T.,€Shenaut,€M.A.,€Schmidt,€M.E.,€Myers,€Jr€O.€¼à0..à(1991).€Soybean€SDS€variety€testing€in€Illinois.€1987„¼1991.1991€Agron.€Abstr.€p94.Ì24.€Gibson,€P.T.,€Shenaut,€M.A.,€Njiti,€V.N.,€Suttner,€R.J.€&€¼à0..àTaylor,€D.€(1993).€Soybean€SDS€yield€effects€in€varietyÏtrials.€1993€Agron.€Abstr.€p86.ÌGingera,€G.R.,€Davis,€D.W.€and€Groth,€J.V.€1994.€PedigreeÏselection€for€improved€partial€resistance€to€common€leaf€rust€inÏsweet€corn.€Crop€Sci.€34:615„Ì25.€Guo,€M.,€Lightfoot,€D.A.,€Mok,€M.C.,€Mok,€D.W.S.€(1991).€¼à0..àAnalyses€of€òòPhaseolusóó€òòvulgarisóó€L.€and€òòP.coccineusóó€Lam.Ïhybrids€by€RFLP:€preferential€transmission€of€òòP.vulgarisóóÏalleles.€Theor.€Appl.€Genet.€81:703„709.Ì27.€Hartwig,€E.E.€&€Epps,€J.M.€(1973).€Registration€of€Forrest€¼à0..àsoybeans.€Crop€Sci.13:287.ÌJung,€C.,€M.€Kleine,€F.€Fischer€and€R.G.€Herrmann.€1990.€AnalysisÏof€DNA€from€a€òòBeta€procumbensóó€chromosome€fragment€in€sugar€beetÏcarrying€a€gene€for€nematode€resistance.€Theor.€Appl.€Genet.Ï79:663„672.ÌÌKari,€A.G.,€Griffiths,€E.€1993.€Components€of€partial€resistanceÏof€barley€to€òòRynchosporium€secalisóó:€Use€of€seedling€tests€toÏpredict€field€resistance.€Anal.€Appl.€Biol.€123:545„€€Ì35.€Keim,€P.,€Shoemaker,€R.C.,€Palmer,€R.G.€(1989).€Restriction€¼à0..àlength€polymorphism€diversity€in€soybean.€Theor.€Appl.ÏGenet.€77:786„792.ÌMartin,€G.B.,€Williams,€J.G.K.€and€Tanksley,€S.D.€1991.€RapidÏidentifiecation€of€markers€linked€to€a€òòPseudomonasóó€resistanceÏgene€in€tomato€using€random€primers€and€near„isogenic€lines.ÏProc.€Natl.€Acad.€Sci.€USA€88:2336„2340.Ì51.€Matthews,€W.J.,€Njiti,€V.N.,€Gibson,€P.T.,€Shenaut,€M.A.€¼à0..à(1991).€Inheritance€of€soybean€SDS€response€in€segregatingÏF5€and€F6€derived€lines.€Soybean€Genetics€Newsletter.Ì52.€McClean,€K.S.€&€Lawrence€G.W.€(1993).€Localized€influence€of€òò¼à0..àHeterodera€glycinesóó€on€sudden€death€syndrome€of€soybean.ÏJournal€of€Nematology€25:674„678.ÌMesseguer,€R.M.,€Ganal,€M..,€de€Vicente,€M.C.,€Young,€N.D.ÏBolkan,€H.€€and€Tanksley,S.D.€1991.€High€resolution€RFLP€mapÏaround€the€root„knot€nematode€resistance€gene€(òòMióó)€in€tomato.ÏTheor€Appl.€Genet.€82:529„536.ÌÌ53.€Michelmore,€R.W.,€Paran,€I.,€Kesseli,€R.V.€(1991).ÏIdentification€of€markers€linked€to€disease„resistance€genes€byÏbulked€segregant€analysis:€A€rapid€method€to€detect€markers€inÏspecific€genomic€regions€using€segregating€populations.€Proc.ÏNatl.€Acad.€Sci.€88:9828„9832.ÌÌMiklas,€P.N.€and€Grafton,€K.F.€1992.€Inheritance€of€partialÏresistance€to€white€mold€in€inbred€populations€of€dry€bean.€CropÏScience€32:943„Ì55.€Mulrooney,€R.P.€(1988).€Soybean€disease€loss€estimates€for€¼à0..àsouthern€United€States€in€1987.€Plant€Dis.€72:915.Ì56.€Myers,€O.,€Schmidt,€M.€(1988).€Registration€of€the€Pyramid€¼à0..àsoybean.€Crop€Sci.€28:375„376.Ì57.€Myers,€O.,€Rombach,€J.,€Buettner,€M.,€Schmidt,€M.€&€Gibson,ÏP.€à0..à(1990)€Selection€methods€for€identifying€high€yield€inÏsegregating€cyst€nematode€resistant€soybean€populations.Ï1990€Agron.€Abstr.€p102.Ì58.€Myers,€G.O.,€Anand,€S.C.€(1991)€Inheritance€of€resistance€andÏà0..àgenetic€relationships€among€soybean€plant€introductions€toÏraces€of€soybean€cyst€nematode.€Euphytica€55:197„201.ÌNewton,€A.C.€and€Thomas,€W.T.B.€1993.€Evaluation€of€sources€ofÏpartial€resistance€to€mildew€in€barley€using€enzyme„linkedÏimmunosorbent€assay€and€other€assessment€methods.€EuphyticaÏ66:27„Ì59.€Nickell,€C.D.,€Schwenk,€F.W.€&€Schapaugh,€W.T.€(1981).€à0Ð ..Ð 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àsyndrome€of€soybean€in€Iowa.€Plant€Dis.€78:830ÌÌYoung,€N.D.,€Manacio„Hautea,€D.,€Danesh,€N.S.€and€Kumar,€L.€1994.ÏMapping€oligogenic€resistance€to€powdery€mildew€in€mungbean€withÏRFLPs.€Theor€Appl.€Genet.€84:839„844.ÌÌYu,€Z.H.,€Mackill,€J.M.,€Bonham,€J.M.€and€Tanksley€S.D.€1991.ÏTagging€genes€for€blast€resistance€in€rice€via€linkage€to€RFLPÏmarkers.€Theor.€Appl.€Genet.€84:839„844.€Ì€Ì100.€Zhang,€H.S.,€De€La€Rosa,€R.,€Niks,€R.E.€(1994).€Role€ofÏpartial€resistance€to€òòPuccinia€hordeióó€in€barley€in€the€defense€ofÏbarley€to€inappropriate€rust€fungi.€Physiological€and€MolecularÏPlant€Pathology€45:219„226.ÌÌÌ€Nickell€et€al€1994).€€ÌÌÇ€€€€€òòEóó.òòcolióó€cultures,€stored€in€„70òòoóóC€freezer,€were€grown€in€50Ïml€LB€Broth€(40€ððg/ml€Ampicillin,€1%€NaCl,€1%€Bacto„Tryptone,Ï0.5%€Bacto„Yeast€Extract)€in€a€Lab„line€Orbital€incubator€atÏ37òòoóóC.€Plasmid€DNA€was€extracted€and€purified€€using€PromegaÏWizard€Miniprep€and€QIAprep„spin€plasmid€kit€(Qiagen).ÌÌLINKAGE€GROUP€AÌÌA117aà` ..` àภ..¸ àRhg4à..ààh..hààÀ..Ààà..ààp..pàK636cà .. ààx..xàK636dÌòò€|€€€€óó//òò€€€€€|€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€|€€€€€€€€€€€|€€óó€ÌÌ|SDS|à` ..` à€€|SCN|€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€|„SDS„|ÌÌÌÌLINKAGE€GROUP€BÌà..àA847a,bÌòò€€€€€€€|€€€€€€€€óóÌ€€€€Ì€€€€|„SDS„|Ì€€€€|„SCN„|ÌÌÌÌLINKAGE€GROUP€CÌÌOPO05d?€€€OPO14fà..à€€€€K455a,cà..à€€€€€€€€OPO05e€€€€€€€€ÏOPF05g?à` ..` àÌòò€€|€€€€€€€€€|€€€€€€€€€€€|€€€€€€€€€€€€€€€|€€€€€€€€€€€€€€|€€€€€óó€Ì€€€€€Ì|„YLD„|€€€€€€€€€€€€€€|„YLD„|Ì€€€€€€€€€€€€€€€€€€€€€|„„„„„„„„„SDS„„„„„„„„„„|€€€Ì|„SCN„|€€€€€€€€€€€€€€|„„„„„„„„„„„„„„„„„SCN„„„„„„„„„„„„„„„„„|ÌÌÌÌLINKAGE€GROUP€DÌÌSOPF20eà` ..` àOPC01m€€€€€€€€€K411a,bÌòò€€€|€€€€€€€€€€|€€€€€€€€€€€€€€|€€€€€óóÌÌ|„„„„„„R6„„„„„„„|ÌÌÌÌFigure€7.€€Proposed€linkage€groups,€A,€B,€C,€and€D,€showingÏpositions€of€à¸ ..¸ àà..àloci€identifying€regions€of€DNA€associatedÏwith€SDS€à` ..` àภ..¸ àresponse,€SCN€resistance,€yield€under€SDSÏpressure,€and€R6€à..ààh..hàmaturity€date.€€ÌÇLINKAGE€GROUP€FÌà..àÌA020à..àà` ..` àภ..¸ à€€€€€A112b,cÌòò€€|€€€€€€€€€€€€€€€€€€€€€€€|€€€óóÌ€€€€€€€€€€€€€€€€€€€€€€€|„„R6„|à..à€€€€€€ÌÌÌÌLINKAGE€GROUP€GÌà..àÌA186bà` ..` àภ..¸ àK644aàh..hààÀ..ÀàA186aàp..pààÈ..ÈàK644bÌòò€€|€€€€€€€€€€€|€€€€€€€€€€€|€€€€€€€€€€€|€€€€€óóÌ€€€€€€€Ì€€€€€€€€€€€€€€€€€€€€€€€|„SDS„|Ì€€€€€€€€€€€€€€€€€€€€€€€|„YLD„|Ì|„SCN„|€€€€€€€€€€€€€€€€|„SCN„|ÌÌÌÌLINKAGE€GROUP€LÌÌK385aà` ..` àภ..¸ à€€€€K385b€€OPF13bÌòò€€|€€€€€€€€€€€€€€€€€||€€€€€€€€óóÌà..à€€€Ì€€€€€€€€€€€€€€€€€€€€€|„„R6„|ÌÌÌÌLINKAGE€GROUP€NÌÌSOPC04c€€SOPO04i€€OPF04e€€€OPC01kà..à€A071c€€€€SOPC11hàx..xààÐ ..Ð àÏA071aÌòò€€€|€óó//òò€€€€|€€€€€€€€€|€€€€€€€€|€€€€€€€€|€€€€€€€€€|€€€€€€€€€€€€|€€óóÌ€€€€Ì€€€€€€€€€€€€€€€€€€€|„„„„SDS„„„„|Ì€€€€€€€€€€€€€€€€€€€|„„„„YLD„„„„|Ì€€€€€€€€€€€€€€€€€€€€€€€€€€€€€|„„„„„„„„„SCN„„„„„„„„„|Ì€€€€€€€€€€€€|„„„„„„„„„R6„„„„„„„„|€€€€€€€ÌÌÌÌÌFigure€8.€€Proposed€linkage€groups,€F,€G,€L,€and€N,€showingÏpositions€of€à¸ ..¸ àà..àloci€identifying€regions€of€DNA€associatedÏwith€SDS€à` ..` àภ..¸ àresponse,€SCN€resistance,€yield€under€SDSÏpressure,€and€R6€à..ààh..hàmaturity€date.€€ÌÇÌ1.9€€€€€€€€€€€€1.0€€€1.0€€€€1.6€€€€€€16.2€€€€2.5€€€€€€LOD€ROOTÏSCNÌÌ1.9€€€€€€€€€€€€1.0€€€1.0€€€€1.7€€€€€€11.3€€€€2.6€€€€€€€€LOD€SCNÌ€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€rhgÌOPO05d€€€€€€€€K455€€OPO05e€€€€€€€€€€€€€|€€€€€OPF05gÌ„|„„„„„„„„„„„„„|„„„„„|„„„„„„„|„„„„„„„„„|„„„„„„|„„„„„„Ì€€€€€€€€€€€€€€€€€€€€€€€€€€€€€|Ì€€€€€€€€€€€€€€€€€€€€€€€€€€€€rsdÌ0.7€€€€€€€€€€€2.45€€€2.62€€€2.78€€€€€€€2.42€€€1.98€€€€€€LOD€DIÌÌÌÌÌ0.02€€€€€€€0.12€€€€€€€€€€0.94€€€€€€€€€€1.22€€€€€€€€€€€1.24€€€€€Ï12.8€€€€€€0.90€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€Ï€€€€€€€€€€€rhgÌSOPO04i€€€OPF04e€€€€€€€OPC01k€€€€€€€€€A71c€€€€€€€€SOPC11h€€€€€€|€€Ï€€€€€A71aÌ„|„„„„„„„„„|„„„„„„„|„„„„„|„„„„„„„„„„„„„|„„„„„„„„„„„„„|„„„„„„„„„|„¼„„„„„„„„„|Ì€€€€€€€€€€€€€€€€€€€|Ì€€€€€€€€€€€€€€€€€€rsdÌ0.48€€€€€€€1.33€€€2.05€€1.80€€€€€€€€€€€1.80€€€€€€€€€€0.57€€€€€Ï1.19€€€€€€€1.04ÌÌÌÌ4.5€€€10.6€€€€0.66€€€1.5Ì€€€€€€rhgÌK644a€€|€€€€€A186a€€K644bÌ„|„„„„„|„„„„„„„|„„„„„|„„„„ÌÌÌ0.90€€0.88€€€€0.46€€€0.90ÌÌÌÌÌ0.26€€€€€€15.1€€€€€€€€€€€€€€€€€€€€€€€€€14.2€€€€€€€€€€€0.6ÌA20€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€€A112Ì„|„„„„„„„„„|„„„„„„„„„„„„„//„„„„„„„„„„„„„|„„„|„„„„„„„„„„|„ÌÌ0.65€€€€€€2.59€€€€€€€€€€€€€€€€€€€€€€€€€€€€2.61€€€€€€€1.2ÇÔ€XXXÔÔU€1*Ͼ¦ `(CG TimesScalableUÔÌÌÌÌNovember€21,€1994ÌÌDear€Nix,ÌÌI've€reviewed€the€materials€and€methods€and€they€are€OK€if€a€little€long€and€wordy€andÏlacking€references.€How€are€the€figures€coming€could€you€fax€drafts€so€I€can€write€theÏresults€text€etc.ÌÌ€€€€€Attached€are€the€results€from€mapmaker€QTL.€The€only€trick€you€missed€was€changingÏthe€header€on€the€data€file€to€F2€backcross€after€Mapmaker€EXP3.0!€€I€tested€Lgs€C,€D,€F,ÏG,€L€and€N€for€all€five€traits,€SCN,€Maturity,€Yield,€DI€and€DS€and€found€very€fewÏsignificant€intervals.€None€for€yield€or€maturity,€1€for€DS€(lgC),€3€for€DI€(lgC,€N,€F!)€andÏfour€for€SCN€(lg€C,N,€F,€G).SDS€intervals€were€barely€significant€(LOD€2.0„2.8)€andÏexplained€little€variability€(10„20%).€SCN€intervals€were€strong€(LOD€10„16)€and€eachÏexplained€almost€all€the€variability!€I€assumed€the€bimodal€distribution€of€the€SCN€trait€foolsÏQTL€somewhat€but€square„root€transformation€of€the€SCN€index€didn't€help€much€€and€bothÏYoung€and€Webb€used€untransformed€data€too€so€I€guess€its€OK.€In€any€case€in€the€paper€weÏshould€stress€the€lack€of€linkage€between€the€seven€loci€and€linkage€within€the€best€3€toÏvalidate€the€rest€ÌÌ€€€€€Did€you€receive€the€Kinkos€refund€yet?€The€Teachers€travel€refund€is€still€being€typed,Ïthis€time€as€an€invoice€voucher,€maybe€by€Xmas!ÌÌHappy€Thanksgiving,€dont€eat€too€much€turkeyÌÌÌSincerelyÌÌÌDavid€A.€LightfootÌAssistant€ProfessorÌÌPS.€I€just€heard€from€Kerrie€that€she€is€pregnant€for€June!ÇÔ€XXXÔÔO€.<þ6X9`("Courier 10cpiOÔÌÌÌÌÌAugust€9,€1995ÌÌDr.€Perry€B.€Cregan,ÌUSDA„ARS,ÌSoybean€and€Alfalfa€Research€Lab.,ÌBldg.€011,€HH„19,ÌBARC„West,ÌBeltsville,€ÌMD20705„2350.ÌÌÌDear€Perry,ÌÌ€€€€€€€€please€find€enclosed€a€revised€copy€and€disc€of€aÏmanuscript€for€Crop€Sciences€as€part€of€the€"C7"€experiment€on:€Ìò ò€€€Genetic€Mapping€of€Loci€underlying€Field€Resistance€to€SoybeanÏà..àSudden€Death€Syndrome€(SDS).Ìó óby€€€€€€€N.€Hnetkovsky,€S.€Chang,€T.W.€Doubler,Ì€€€€€€€€€€€P.T.€Gibson€and€D.A.€Lightfootòò*óó.ÌÌI€appreciated€the€points€you€brought€up€and€added€a€fewÏalterations.€I€enclosed€the€marked€copy€of€the€original€aÏrevision,€disc€copy€and€a€letter€referring€to€your€letter€pointÏby€point.€€€€€€€ÌÌSincerely,ÌÌÌDavid€A.€LightfootÌAssistant€ProfessorÌÇ1.€Added€E„mail€address.ÌÌ2.€Added€Scientific€names.ÌÌ3.€The€third€population€was€Ripley€x€Spencer€the€first€two€wereÏEssex€x€Forrest€and€Pyramid€x€Douglas.€I€hope€the€text€is€nowÏclearer.ÌÌ4.€As€noted€in€the€text€C91€and€C92€were€separate€fields€(about€2Ïmiles€apart)€so€using€locations€would€still€be€justified€althoughÏthis€is€closer€than€any€other€pair€of€locations.€With€a€patchyÏdisease€like€SDS€using€environment€would€imply€a€uniformityÏwithin€fields€that€does€not€exist.ÌÌ5.€It€does,€although€intrinsically€repetitious€we€included€theÏnumber€of€reps€to€clarify€a€point€two€of€the€reviewers€broughtÏup.€Given€the€variety€of€scientists€likely€to€read€the€paper€IÏwould€like€to€retain€this€for€clarity.ÌÌ6.€Feet€converted€to€meters.€Ooops€sorry.ÌÌ7.€Adjusted€as€noted.€I€felt€uncomfortable€with€the€form€of€theÏreference€however,€perhaps€adding€Spechts€recent€paper€as€aÏreference€might€be€better,€please€advise€if€changes€are€needed.ÌÌ8.€A€confusion€arises€over€probes€tested,€probes€actually€usedÏand€loci€scored.€I€think€I€have€clarified€the€point.ÌÌ9.€Interaction€was€the€wrong€term,€recombination€is€correct€(weÏhave€fully€tested€this€now).ÌÌ10.€Actually€the€LSD€was€calculated€Excluding€the€8€reps€perÏlocation€of€Forrest€otherwise€the€LSD€would€have€been€smaller.ÏThis€is€noted€in€the€text.ÌÌ11.€Clarified€as€suggested.ÌÌ12.€Reworded€for€clarity.ÌÌ13.€Yield€as€a€result€of€SDS€DI€resistance.ÌÌ14.€Altered€as€suggested.ÌÌ15.€Better€title.ÌÌ16.€Done