The apple genome: origin of the species and perspectives
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1 The apple genome: origin of the species and perspectives Riccardo Velasco, FEM-IASMA, San Michele all Adige, Trento Bolzano, 6 novembre 2010
2 Apple genome sequencing Cultivar Golden delicious (most important world and Trentino cultivar) Diploid (2n = 34) Genome size: ca. 750 Mb Sequenced 17x (Roche ca. 13x + Sanger ca. 4x) Ca. 70% of genome anchored on linkage map, including ca. 90% of genes. Estimated ca. 2 millions molecular markers (SNPs single nucleotide polymorphisms)
3 Genome sequencing: neverending puzzle sequencing 2 kb Clones Contigs Metacontigs da 2-15 kb Clones BACs e Fosmids Metacontigs da BACs and Fosmids Molecular markers Chromosomes
4 Details on the Golden genome Third round of assembly (15.4x Golden Delicious + 1.5x DH) 122,146 contigs assembled into 20,738 clusters after corrections for overlaps of which 19,084 singletons and 1,654 metacontigs (average lenght ca. 7 kb), for a total length of 658,7 Mb repeat estimation length 80 Mb 1,654 metacontigs are assembled into 605 Mb, out of them 554 cover 535 Mb anchored (max length Mb - from 0 to 31.1 cm of LG15) , 400 TC from 255,000 ESTs 23,058 TC aligned to the genome with 90% of identity and 80% of length - approx. 57,000 genes predicted. - Ca. 93% of predicted genes are positioned in anchored metacontigs # c o n ti g N100 N90 N80 N70 N60 N50
5 2 1 negative control 3 The apple genome: milestones in plant evolution 4
6 Apple applied genetics and genomics Germplasm collection at FEM IASMA Genetic distance between Malus species and the origin of the cultivated apple 23 genes re-sequenced 74 Malus accessions 12 M. x domestica 10 M. sieversii 21 M. sylvestris Splits-Tree (Hamming distance)
7 Apple applied genetics and genomics Cultivar fingerprinting Internally validated system on: 132 apple cv Ca. 200 SNP on the 17 chromosomes 100% of distinguibility between cultivars MDFiesta MDFreedom MDKent MDCoxOrange MDIngridMarie MDDelcoros MDJamesGrieve MDLordLamboune MDAmorosa MDClivia MDPilot MDReanda MDRewenda MDBenDavis MDLaxtonSuperba MDMcIntosh MDWagener MDJanamac MDAkane MDWorcesterPearmain MDny5822McIntosh MDBmypol MDEnterprise MDSpartan MDRenettaBianca MDRenettaGrigiaGranFaje MDCalvilla MDBoskoop MDJonafree MDJonathan MDIdared MDRedRome MDRomeBeauty MDDiwa faw MDEarlychief MDHapke MDJeromine MDOregonSpur MDStarkingDelicious MDCatarina MDFuiji MDCarlaOpaca MDStayman MDBraeburn MDGloster MDCrimson Snow MDFlorina MDPriscilla MDAmbrosia MDGotha MDGrannySmith MDForlady MDBoujade MDDalinred MDAntonovka MDGravenstein MDDiscovery MDSanLugano MDPiros MDRenettaAnanas MDRosmarinaBianca MDSpitzlederer MDNapoleone MDDurelloDiForli MDRosaDiCaldaro MDSila MDGenevaCrab MDGoldenHornet MDRedField MDAnnurca MDPioneerScarlet MDRadiant MDWinterGold MDRedJade MDVanEsentine MDGoldenGem MDJohnDownie MDPearmainDorata MDEvereste MDPlatycarpa MDDolgo MDMelaDellaSerla MDResista MDSunkrisp MDRedEarly MDEarlyRedGold MDGala MDRubicon MDRubyGala MDInitialPajam2 MDSonya MDSansa MDModà MDRubin MDTopaz MDRosana MDGoldenOrange MDElstar MDRubens MDElan MDGoldenDel MDMutsu MDPinkGold MDDelblush MDGoldrush MDRubinette MDDoriane MDPinkLady MDLygol MDPinova MDGingerGold MDRebella MDPrima MDRubinola MDRetina MDBrina MDRenora MDSummerfree MDPri14126 MDStettino MDBelfioreGiallo MDSaturn MDCoop36 MDDelgrared MDCameo MDPrimiera MDAriwa MDDelorina MDAmbassy MDTunda MDEarlyGold MDMonroe Red Del clones Gala clones cv Initial (Gala x Redfree) GD clones Micheletti et al. unpublished Least difference between cvs Difference between apple clones ~ experimental error
8 Golden Delicious Scarlet IASMA reference map Golden delicious Scarlet spur 270 individuals 1,571 markers 1,413 SNP 158 SSR Map length: 1,159.7 cm Average marker interval: 0.7 cm Troggio et al. submitted
9 Linkage based-qtl mapping projects at FEM-IASMA Mapping populations for genome anchoring and generation of consensus map Golden Delicious x Scarlet (reference cross, 270 ind.) (quality) Golden Delicious x Braeburn (96 ind.) (quality) Golden Delicious x Freedom (300 ind.) (mapping Va -scab resistance) Mapping populations Projects for gene/qtl mapping (Internal + collaborations) Fuji x Pink Lady (96, quality, internal project) Fuji x Delearly (96, quality, internal project) Golden Delicious x M. hupehensis (Erwinia, internal project) Golden Delicious x M. fusca (Erwinia, flowering biology, internal project) Golden Delicious x M. prunifolia (Erwinia, internal project) GoldRush x McIntosh Wicjik (Habitus, internal project) Golden Delicious x McIntosh Wicjik (103, Habitus, internal project) Gala x X-3191 (self-thinning, coll INRA) Pink Lady x Idared (192, self-thinning, internal project) Gala x Rubinette (96, resistance, quality, internal project) Mapping populations collaborations Royal Gala x Braeburn (IRTA-HortResearch-INRA) TN x Discovery (INRA) Gala x Elstar (INRA) Pyrus communis Max Red Burtlett x P. pyrifolia (HortResearch)
10 Apple applied genetics and genomics QTL analysis for skin color Golden delicious Scarlet spur 270 F 1 plants 2-year phenotype 705 markers QTL analysis (MapQTL6, CP, IM) 120 LOD GD_SNP00332 GD_SNP01200 GD_SNP00352 GD_SNP00886 GD_SNP00766 GD_SNP00205 GD_SNP01817 GD_SNP01416 CN GD_SNP00659 GD_SNP01066 GD_SNP00206 GD_SNP02482 GD_SNP01596 CH01h02 GD_SNP01466 HI01d01 GD_SNP00509 GD_SNP02441 GD_SNP02100 GD_SNP02014 GD_SNP00331 GD_SNP02046 GD_SNP00109 HI23d06 GD_SNP01959 CH05c07 GD_SNP01880 GD_SNP00947 GD_SNP02031 GD_SNP00400 CH01f03b GD_SNP01529 GD_SNP00337 GD_SNP01700 GD_SNP00893 GD_SNP01367 GD_SNP01706 GD_SNP00280 GD_SNP00514 GD_SNP01911 GD_SNP01948 GD_SNP01771 GD_SNP00169 GD_SNP02564 GD_SNP01646 GD_SNP Chromosome LOD Genotypic mean value Golden1/Scarlet(+) Golden1/Scarlet(-) Golden2/Scarlet(+) Golden2/Scarlet(-) Rf
11 Apple applied genetics and genomics QTL analysis for self-thinning Apple overproduction is currently controlled by chemical thinning (to be strongly restricted for health and environmental risks and cost concerns) Sources of genetic variability is available QTL analyses on crosses: R. Gala x INRA X-3191 Pink Lady x Idared LOD Zini et al. unpublished Chrom. 3 linkage map
12 Apple applied genetics and genomics Cloning of the Co gene Golden Delicious x McIntosh Wijick 1,298 plants screened (markers and phenotype) Ca. 1,800 new seeds in 2009, now in pots Locus lower to < 1 cm, ca. 300 kb, 3-4 candidate genes. Chr10 600,000 bp interval
13 Breeding by design Example of gene/qtl pyramiding
14 Molecular markers assisted selection n GD traditionsl x Breeding by design assisted M. floribunda Source of Vf gene, resitance to apple scab
15 Acknowledgements FEM - Istituto Agrario di San Michele all Adige Sequencing/Mapping Silvio Salvi, Michela Troggio, Massimo Pindo, Diego Micheletti, Giuseppina Coppola, Fabrizio Costa, Paolo Baldi, Mickael Malnoy, Francesco Salamini, Roberto Viola Bioinformatics Paolo Fontana, Alessandro Cestaro, Marco Moretto, Stefano Toppo Annotation Claudio Moser, Giulia Malacarne, Azeddine Si-Ammour Breeding Pierluigi Magnago, Marco Fontanari Dep. of Plant System Biology, Gent University, Belgium Yves Van de Peer, Jeffrey Fawcett, Lieven Sterck, Klaas Vandepoele Roche/454 Egholm Michael, Jason Affourtit, Tim Harkins, Kodira Chinnappa Amplicon Express Amy Mraz, Keith Stormo, Quanzhou Tao, Robert Bogden Myriad Genetics Andrey Zharkikh, Alexander Gutin, Dmitry Pruss, Satish Bhatnagar, Natalia Gutin, Glenn Eldredge, Jerry Lanchbury, Mark Skolnick, Dustin Cartwright Washington State University Amit Dhingra INRA - Charles-Eric Durel, Yves Lespinasse, Pauline Lasserre University of Western Cape, South Africa Jasper Rees HortResearch Sue Gardiner, Andrew Allan, Roger Hellens, Vincent Bus, David Chagne
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