16 apple germplasm strcture and tools for germplasm curators durel charles eric

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Information about 16 apple germplasm strcture and tools for germplasm curators durel...

Published on February 28, 2014

Author: fruitbreedomics

Source: slideshare.net

Genetic structure of European apple germplasm Molecular markers as tools to manage practical issues in germplasm collections C.E. DUREL INRA, Angers

Major questions when curating a germplasm collection : • Is my accession corresponding to the true genotype ? (TTT = « True To Type ») • Is my accession unique or redundant within my collection or with other collections ? • Are these 2 accessions related ? • Are these 2 accessions genetically close or distant ? • How representative of the genetic diversity is my collection ? • Is my collection structured in subgroups ?

Markers available in apple (isozymes, RAPD, AFLP) SSR : (Simple Sequence Repeats) > 660 published http://www.hidras.unimi.it/ SNP : (Single Nucleotide Polymorphism) 8k / 18k / … 420k GBS : (Genotyping by Sequencing) (Elshire et al., 2011)

Fingerprinting SSR SSR1 SSR2 SSR3 SSR4 90-100 120-126 164-168 205-209 96-100 SNP2 98-102 AG 90-100 126-132 SNP3 122-130 CC 120-126 164-170 SNP4 162-170 GT 164-168 205-211 … 207-213 AA AG CT GG vr C CC AA TT TT vr D AC AG CC GT Cvrs Cvr A SNP Cvr B SNP1 vrs Cvr C vr A Cvr D AC vr B … … The more markers, the better … i.e., the more specific the fingerprint is. 205-209 Genetic fingerprinting

Genetic distances SSR SSR1 SSR2 SSR3 SSR4 Cvr A 90-100 120-126 164-168 205-209 Cvr B 96-100 126-132 164-170 205-211 Cvr C 98-102 122-130 162-170 207-213 Cvr D 90-100 120-126 164-168 205-209 Cvrs  Genetic distances (dissimilarities) according to common/distinct marker profiles Cvrs Golden B Gala Braeburn CG 10 YD Cvrs Cvr A 0 Cvr B 0.5 0 Cvr C 1 0.87 0 Cvr D 0 0.5 1 0

Is my accession True To Type ? SSR SSR1 SSR2 SSR3 SSR4 Cvr A 90-100 120-126 164-168 205-209 Cvr B 96-100 126-132 164-170 205-211 Cvr C 98-102 122-130 162-170 207-213 Cvr D 90-100 120-126 164-168 205-209 Cvrs Clones (+/- mutations) Same fingerprints dij = 0

Disctinctness SSR SSR1 SSR2 SSR3 SSR4 Cvr A 90-100 120-126 164-168 205-209 Cvr B 96-100 126-132 164-170 205-211 Cvr C 98-102 122-130 162-170 207-213 Cvr D 90-100 120-126 164-168 205-209 Cvrs ~ Unrelated ~ No matching alleles dij ~ 0.0xx

Triploidy checking N° echantillon Diploid / Triploid CH01f03b CH01h01 CH02c06 CH02d08 CH04e05 CH05f06 NZ05g08 0001 Bédange de Nantes Diploid 174/186 121/127 254/256 215/258 205/205 171/185 127/163 0002 Belle de fumée Triploid 141/174 119/123/127 232/242/254 215/229 178/213 179/189 127/127 0003 Belle fille du Penthièvre Triploid 141/162/186 119/121/127 - 229/258 178/205/213 179/187/189 133/149 0004 Blanc d'été Diploid 141/141 125/127 254/254 211/231 205/205 185/189 127/142 0005 Bercelien Triploid 162/174/186 125/137 232/254 215/258 178/213/219 171/181/187 127/129 0006 Boblin Diploid 141/174 105/137 232/262 217/260 178/219 177/189 121/121 0009 Charles pitrel Diploid 174/186 133/137 254/254 258/258 178/219 179/185 129/142 0010 Coaquin Triploid 141/174 125/133 234/238/252 215/229/258 178/205/219 171/183/185 144/151/163 0011 Cul d'oie Diploid 141/141 127/137 232/246 209/229 178/205 177/181 127/127 0012 Cul na Diploid 141/174 119/133 220/232 215/256 178/222 179/181 127/127 0013 Cyriac Diploid 141/186 137/137 232/232 215/229 178/219 181/187 127/127 … Accessions with 3 alleles per marker  triploid apple varieties

Is my accession unique or redundant within my collection or with other collections ? duplicate 2n 69 genotypes (88 accessions) Collection 1 (408 DNA samples received) unique 2n 190 genotypes (209 accessions) Duplicates - Belgium 8 7 6 5 4 3 2 1 0 1 6 11 16 21 26 31 36 41 46 51 56 61 66 71 76 81 86 91 96 101 106 unique 3n 59 genotypes (79 accessions) duplicate 3n 10 genotypes (16 accessions) Belgium Czech_Republic France Italy Sweden United_Kingdom

Are these 2 accessions related ? http://en.wikipedia.org/wiki/ File:Pedigree_marker_information.jpg Large data set  Relatedness coefficient (ML-Relate software ; Kalinowski et al., 2006) Parent-Offspring (0.5) / Full-sibs (0.25) / Half-sibs (0.125) / …

Parentage analysis Parentage inference conditionally to the available marker data: Female parent ? Male parent ? Correct marker allele inheritance ? Known variety  Software CERVUS (Kalinowsky et al., 2005)

Parentage analysis Inference of unknown parents: Pomme raisin Full-Sibs: Rose de Berne Requirements: - very large data set - >= 20 SSR Calville rouge d’hiver Rose d’Ajoie Blaser (2 Swiss cvrs) Caution !: - Are the accessions TTT ?? - The true cross could derive from mutants of the identified parents Lassois et al. (in prep.)

Global relatedness Putative relatedness tree in grape: IBD computed on >5300 SNPs (Myles et al., 2010)

Is my collection structured in subgroups ? FRB-funded project Cider Dessert - Old Dessert - New Lassois et al. (in prep.)

Is my collection structured in subgroups ? Wild apples Domesticated apple

Is my collection structured in subgroups ? FruitBreedomics – WP4 : 2750 accessions fingerprinted with 16 SSR

Is my collection structured in subgroups ? Faint structure in 3 European regions West North-East South (1550 diploid genotypes with known geographic origin)

How representative of the genetic diversity is my collection ? Collection 1

How representative of the genetic diversity is my collection ? Collection 2

Can I get a subset of cvrs representative of my (neutral) genetic diversity ? Core collection 97% 100 Captured allelic richness (%) 80 60 40 20 0 0 100 200 300 400 500 600 N° of accessions of the core Lassois et al. (in prep.)

Technical issue for SSR : ajusting allele sizes  choice of genotypes covering almost all SSR alleles 96 accessions covering almost all SSR alleles  Facilitating the alignment of SSR alleles

Limits Marker versus Phenotype mutation Marker mutation rate = 10-4 for SSR 10-6 for SNP Phenotypic mutations : Colour Architecture Acidity Vigor …     Gala, Red Delicious McIntosh vs Wijick Usterapfel LA vs HA M9 rootstock Genetic mutation Regulation modification Cell layers L1/L2/L3

SSR marker mutation Marker CH03d12 Des moissons 1 single allelic difference over 24 markers Pomme Livre Allelic mutation Belle Louronnaise Demie sûre

Markers can help germplasm curators • • • Quality insurance : Grafting/labelling error Erroneous pomological identification Information about duplicates within and between collections (but not mutants !) Possible inferences of parentage (when enough markers and large data set)  looking backwards how selection has been done empirically by farmers and gardeners • Representativeness/specificity of the collection  To be combined with phenotypic data • Optimal choice for core collection • Genome-Wide Association Studies

Technical and financial issues Multiplexing Analysis Cost Information SSR : Low Long Low High (multi-allelic) SNP : High Short GBS : High Long (bioinformatic) High 8k ~ 50 € 18k ~ 65 € 420k ~ 165 € Low Low / marker but numerous SNP Missing data (imputation)  Major goal : ~500-1000 SNP at 5-10 € ???

Thank you • • • • • • • • • • • • M. Lateur, P. Houben (CRA-W) S. Tartarini, L. Dondini (UNIBO) F. Paprstein, J. Sedlak (RBIPH) M. Ordidge (Reading Univ.) F. Fernandez, K.M. Evans (EMR) H. Nybom, L. Garkava-Gustavsson (SLU) C. Miranda, J. Urrestarazu (Un. Navarra) J. Gassmann (Agroscope) K. Antonius (MTT) I. Suprun (SKZNIISIV, Krasnodar) A. Pikunova (VNIISPK, Orel) C. Denancé, E. Ravon, L. Feugey, A. Guyader, R. Guisnel, L. Lassois (INRA)

Thank you for your attention!

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