The$IWGSC:$$$ Strategies$&$Ac4vi4es$to$Sequence$ the$bread$wheat$genome$

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1 The$IWGSC:$$$ Strategies$&$Ac4vi4es$to$Sequence$ the$bread$wheat$genome$ Kellye$A.$Eversole! IWGSC!Execu,ve!Director! &$ The$IWGSC$ Wheat$Breeding$2014:$$Tools,$Targets,$&$Progress$ Harpenden,$United$Kingdom$ 29$January$2014$ The!Interna,onal!Wheat!Genome!Sequencing! Consor,um! Launched)in)2005)on)the)ini0a0ve)of)Kansas)Growers)) CoMchairs$ R.$Appels$ J.$Dvorak$ C.$Feuillet$ B.$Gill$ B.$Keller$ Y.$Ogihara$ Execu,ve!director! K.$Eversole$ General!members!! Coordina,ng! Commi:ee! Sponsors!

2 Why! Vision! Lay!a!founda,on!to!accelerate!wheat!improvement! Increase!profitability!throughout!the!industry! High!quality!annotated!genome!sequence,! comparable!to!rice!genome!sequence! Physical!mapJbased,!integrated!and!ordered! sequence! $Wheat$Improvement$is$Complex!! Yield potential and yield stability! Adaptation to climate change! Durable resistance to biotic stress! Quality of grain and co-products

3 Training, capacity building -> expertise and critical mass Complexity!Requires!an!Integrated!Toolbox! Improved wheat varieties Managing!the!17!Gb,!Hexaploid!Genome! AA BB Triticum aestivum (2n = 6x = 42) 1C ~ 17,000 Mbp DD Dissection of the genome into single chromosomes (arms) Sheath fluid D Flow chamber Fluorescence emission Laser Excitation light B A Deflection plates Scattered light ; Waste Doležel et al., Chromosome Res. 15: 51, 2007 " Chromosomes: Mbp ( % of the genome) " Chromosome arms: Mbp ( % of the genome) Chromosome genomics! Chromosome specific BAC libraries and sequencing IEB

4 Faster!and!cheaper!methods!to!tackle!the!wheat!genome! h:p://flxlexblog.wordpress.com/2012/12/03/ developmentsjinjnextjgenera,onjsequencingjaj visualisa,on/! We:erstrand!KA.!DNA!Sequencing!Costs:!Data!from!the!NHGRI!! A!sequence!for!whom!&!for!what!purposes! Feuillet)et)al,)TIPS,)2010)

5 BAC!by!BAC!vs!WGS) BAC by BAC (physical map)! TEs Genes 500 kb Whole genome shotgun IWGSC!Strategy!for!a!Reference!Wheat!Genome!Sequence! 1. BAC library construction 2. BAC fingerprinting (HICF/WGP) 3. Contig assembly by FPC/LTC GAATTCTTGGTCAGCAATAAGGACAAAGAA! GAATTCCTCCTGTGCGGCCGTTTTA! GAATTCATGCTCCAGCCAACTCATCCAAAA! GAATTCAACAATATAAATATCACCTAAAGC! GAATTCCATCAGTATGGAGTTTCTTCATGC! GAATTCAAGCTTGAGTATTCTATAGTGTCA! GAATTCAATATGTAGAAATTATAGGATGGC! GAATTCACCGGAAGGGGTTCCGGGTGTTTC! GAATTCCGGAACGCTTCCGGAGACCAAACA! GAATTCTGGAATCACTTGATGATGAATCAG! GAATTCTGGAGTCTTCCTTTCCGCAGCGAA! GAATTCAACGAAATAGTTATGAAATAGAAG! GAATTCTGGGGTAACGATGGAATCCTACAC! GAATTCTGTCAACGACACTATCATCAGGAA! GAATTCTTCGATGCACTGAGTCGCACGGTT! GAATTCTTGTGTGAGGTCCGCGTCCTAGAT! 4. MTP sequencing /Scaffold assembly 5. Pseudomolecule construction (meiotic/ld/rh mapping) 6. Automated and curated annotation

6 Roadmap!to!the!Wheat!Genome!Sequence) Physical$mapping$of$individual$ chromosomes$ Survey$sequencing$of$individual$ chromosomes$ Short term Gene catalog Virtual order Markers MTP$sequencing$ Long term A$reference$sequence$anchored$to$the$gene4c$and$ phenotypic$maps$ Current$status$of$individual$projects$ Physical$ maps$ Illumina$ Survey$ sequences$ PseudoM molecules$

7 IWGSC$Chromosome$Survey$Sequencing$Ini4a4ve! Amplified$ DNAM$sorted$ individual$ chromosomes$ A,$S,$D,$$&$ AB$genomes$ (7)$ ~50X$ Illumina$ sequence$ >30X$ Illumina$ sequence$ Chromosome$ arm$sequence$ assemblies$ Read/assembly$ alignment$to$ chromosome$ arms! Gene$modeling,$ virtual$ordering$ (GZ),$ annota4on,$ func4onal,$ structural$ analyses$ Composi4on$ and$evolu4on$ of$the$21$bread$ Wheat$ Chromosomes$ IWGSC!Chromosome!Survey!Sequencing!J!Summary!!) Summary$ Almost!full!wheat!gene!complement!iden,fied!and! allocated!to!chromosome!arms! On!average,!53%!of!genes!virtually!ordered!along! chromosomes! High!level!of!interJ!and!intrachromosomal!duplica,on! Over!3.5!M!markers!mapped!to!con,gs!(1.3M!wheat! markers!+!2.3m!snps)!j!ssr,!est,!dart,!snp!(90k)! markers...! 13.2!million!SNPs!from!POPSeq!aligned!to!con,gs!

8 IWGSC!CSS!Resources!are!Facilita,ng) New!phylogene,c!analyses!of!wheat!genome! evolu,on!! Defini,on!of!core!and!pan!gene!sets!for! Tri0ceae!by!comparing!diJtetraJ!and!hexaploid! genomes!! HomoeologueJspecific!gene!expression!analyses! Wheat!haplotype!map!!E.!Akhunov!! TILLING!projects!in!T.)durum)and!bread!wheat!! UC,!Davis,!JIC,!Rres! Establishment!of!WheatNet!!UC,!Davis! 3B!SEQuencing!Project! Chr$3B$physical$map$ 1,282!BACJcon,gs! 8,441!BACs! Pool!of!10!BACs! 922!pools!!(Roche!454!GSFLX!Titanium,!8!Kb!MP)!! (~35X)!! Sorted$chr.$3B$ Sanger!!!Illumina!(82X)!! (2*600!bp)! (2*108!bp)!! PE!0.5!kb! Catherine!Feuillet!&!Fred!Choulet! 3B$pseudomolecule$assembly$ and$analyses$

9 An!integrated!and!ordered!3B!reference!sequence!) MetaQTL analysis 3B consensus map (5000 markers) 3B Physical map 3B pseudomolecule MapJbased!cloning!projects!on!3B! 40 genes and QTL mapped on 3B # 4722 markers on 3B consensus map, 3102 in 964 SC (679 Mb =82 % of the sequence) # 13 map-based cloning projects! Disease resistance genes (Sr, Lr, Yr, Stb )! Solid stem (saw fly)! Yield! Drought tolerance! Boron transporter! Flowering time! NUE! Chromosome pairing

10 Cur%s!Pozniak! A$Breeder s$perspec4ve! 3B Physical SSt1 3BL Xgwm114 XBE XPSP3001 SSt1 Xgpw4513 Xgwm340 Xgwm247 Xgwm181 Xgwm114 Xfba217 XBE ctg580 ctg854 Xgwm4703 ctg668 Xwmc274 ctg165 Xgwm247 Xgwm340 Xtam63 Xfba310 Xfba133.2 Xfbb293 Xdarts149 Xgwm181 Xgwm547 Xbarc68.2 Xgpw4513 1$year$ J 1!!EST!marker,!not!closer! J!ascertainment!bias! 3$weeks$ J 12!cosegrega,ng!markers!(SNPs)! J!no!phenotyping!in!the!fields! J!No!ascertainment!bias! $$$Speed$Gene$Discovery:$!!!!!!!!!!!!!!!Towards!Cloning!SSt1! 2.8!cM! Approx.!2.2!Mb! 13.3$

11 Power!of!A!High!Quality!Reference!Sequence! Number!of!QTLs!Cloned! Rice$ Wheat$ Feuillet)et)al,)Trends)in)Plant)Sciences,)2010;)Rey)et)al,)unpublished)update)) Wheat!Sequences!Available!Now) WGS$ WCS$ MTP$ CS$WGS$5x$ (2012)$ A$and$D$genome$$ related$ancestors$ (2013)$ CS$chromosome$ survey$sequences$$ (2013)$ Max!50%!!using!synteny!and!gene,c!mapping!(GZ):!«!virtual!order!»!!CS$3BSEQ$ (2013)$ 94%!real!order! completeness$of$informa4on$ Gold$ standard$

12 $$$$$Applica4ons$ 5X$CS$ Ae.$ tausc hii$ T.$ urartu$ $CS$ CSS$ MTP$ based$(3b)$!«!blind!»!snp!marker! development! x$ x$ x$ x$ Targeted!SNP!marker! development! x$ x$ Conversion!rate!in! genotyping!!for!breeding! 45%$ 95%$ MapJbased!cloning!!,me! (years)! 10$ 10$ 10$ 8$ $2$ Fundamental!studies! $$$$$$$$$$$$$$$limited$to$genes$ $complete$ IWGSC!Projects! The$Annotated$Reference$Sequence$ of$the$bread$wheat$genome$ Pseudomolecules$ (1$of$21$completed)$ Chromosome$Shotgun$Sequences$ and$marker$alignment$ (completed)$ Physical$Maps$with$markers$ (12$of$21$completed)$

13 Cur%s!Pozniak! IWGSC!Resources!in!Breeding! BAC Libraries Physical Maps Wheat Survey Sequence 3B Sequence Unlimited!source!of!DNA! markers!for!mas/genomic! Selec,on! Candidate!genes!for!traits!=! perfect!markers! Reduce!the!,me!and!improve! the!success!of!resolving!qtl!! Discovery!and!exploita,on!of! new!alleles! IWGSC!Resource!Access)!!All!resources!accessible!at!URGI,!Versailles! hjp://wheatmurgi.versailles.inra.fr/seqmrepository/$!!raw!sequence!reads!in!the!sra!!!iwgsc!css!assemblies!available!at!urgi!and!ebi!!!iwgsc!css!assemblies!&!gene!models!integrated!into!ensemblplants! hjp://plants.ensembl.org/tri4cum_aes4vum$ $ 272$ins4tu4ons$ $ 40$countries$ $ 52,000$BLAST$searches$ $ 2.500$downloads$

14 Thank!you!for!your!a:en,on!!