2015 International FruitBreedomics Conference

1. Genome Wide Association Study
of two phenology traits in apple
Hélène Muranty, INRA-Angers, France

2. Acknowledgements
 Conference organizers
 Co-authors
• C. Denancé, D. Leforestier, E. Ravon, A. Guyader, R. Guisnel, L. Feugey, F.
Laurens, C.-E. Durel
• J. Urrestarazu, S. Tartarini, L. Dondini, R. Gregori
• M. Lateur, P. Houben
• J. Sedlak, F. Paprstein
• M. Ordidge
• H. Nybom, L. Garkava-Gustavsson
• M. Troggio, L. Bianco, R. Velasco
• M.C.A.M. Bink, W. Kruijer
 INRA-Clermont-Fd, Gentyane platform: C. Poncet, …

3. Why GWAS ?
• Discover and quantify effects of genomic regions
associated to complex traits
substantially increase resolution by using collections
of unrelated individuals
 candidate genes
markers for selection
origin of favorable alleles Yu and Buckler (2006)

4. many generations
Individual M1 M2 M3 M4 Phenotype
1 2 2 2 1 20
2 1 0 0 0 8
3 2 2 0 0 24
4 1 1 2 1 4
5 1 0 0 0 14
6 1 1 2 1 12
-logp
Chromosomal location
GWAS components
Scoring Detection
Mackay et al (2009)
Very high density genotyping
Affymetrix Axiom_Apple480k array

5. Why phenology traits ?
• target cultivar development / growing season
length in production areas
• develop cultivars able to face climate change
challenges

6. Data available - Material
Country Size
F – INRA (CC) 278
B – CRA-W 229
UK – U. Reading (CC) 294
CZ - RBIPH 178
I – UNIBO (CC) 179
SW – SLU 162
1168
6 collections
old and local dessert apple cvrs

7. Phenotypes
trait scale note
flowering
period
1 (Extremely early) ->
9 (Extremely late)
comparison to
reference cultivars
picking date days from 1st January
picking period 1 (Extremely early) ->
9 (Extremely late)
comparison to
reference cultivars

8. Phenotypic analysis
site Flowering period Picking Date Picking period
# data/gt h² # data/gt h² # data/gt h²
F - INRA 3.0 0.88 3.9 0.96 1.8 0.86
B - CRA-W 4.9 0.88 1 - 4.3 0.87
UK - U. Reading ? - 2.0 0.88 1? -
CZ - RPIPH 5.0 0.85 5.1 0.92
I - UNIBO 7.6 0.84 2.1 0.96 6.6 0.94
SW- SLU 3.0 0.81 2.9 0.98 2.9 0.98
all sites 0.82 0.94 0.89
Heritability of the means
genotypic means adjusted for
• year effects: collection per collection analysis
• (site x year) effects: all collections together
𝑌𝑖𝑗= μ + 𝑦𝑒𝑎𝑟𝑖 + 𝑔𝑗 + 𝑒𝑖𝑗
𝑌𝑖𝑗𝑘= μ + (𝑦𝑒𝑎𝑟𝑖× 𝐿𝑗) + 𝑔 𝑘 + 𝑒𝑖𝑗𝑘

9. NMHom: No Minor
Homozygous
2.4%
Genotypes: quality control and filtering
275K
Additional filtering pipeline
technical replicates (GoldenDel)
biological replicates
Mendelian consistency
mapping progenies
parent-offspring pairs
NMHom: No Minor Homozygous 12K
correct Poly High Resolution 360K
UnexpectedHeterozygosity 11K
criteria from SNP Polisher
visual scoring ~1600 SNP good/poor
logistic regression -> quality prediction
Affymetrix Axiom_Apple480k array
487K
SNP Polisher
Samples: DQC > 0.82 and CallRate > 97%
Unexpected
Heterozygosity
2.8%
correct PHR
73.8%

10. Physical map: present drawbacks
• Scaffold orientation on LG undetermined
• arbitrary 1000bp between scaffolds on LG
• some scaffolds attributed to LG without position
• many SNP (~25%) on scaffolds not attributed to any
LG  LG0 (LG18)
• some SNP from previous arrays not located on the
present physical map  LG20

11. GWAS: Model choice to avoid false positives
Y = µ + SNP + e Y = µ + Q + SNP + e Y = µ + K + SNP + e Y = µ + Q + K + SNP + e

12. Population structure in apple
(3.9 %)
(2.9%)

13. Flowering period
CRA-W INRA NFC
RBIPH SLU UNIBO
SNP + Q + K model

14. Flowering period: advanced model
INRA NFC RBIPH
SNP + Q + K model + SNP cofactors
Extended BIC model selection criteria
MLMM Ségura et al (2012)
LG9
LG9 x 2
LG12
LG11
LG9

15. Flowering period: all collections
SNP + Q + K model
SNP + Q + K model + SNP cofactors
Ext BIC best model
MLMM Ségura et al (2012)

16. Flowering period:
Affx-113839215 : LG9 @ 1293620

17. Picking date
INRA NFC
SLU UNIBO
SNP + Q + K model

18. Picking date: INRA+NFC+SLU+UNIBO

19. Comparison to previously detected regions
trait Chr regions in GWAS
(Mb)
region in QTL analysis
(cM (Mb))
comment reference
Flowering
period
9 1.3 -1.6 0.4 (0.6) Belrène, 2 years Celton et al (2011)
2.5
Allard et al
Eucarpia Fruit 201516.9
Picking date 3 28.6 – 30.3 53.2 (26.0) Braeburn, 3 site-
year comb
Chagné et al (2014)
44.3 75 Discovery Liebhard et al
(2003)
QTL position IC length ~10 cM
74kb
676kb
18kb
222kb
127kb

20. Conclusions & Perspectives
• GWAS can detect already known QTL = proof
of concept
• Variation explained by kinship (+ structure) =
small effect QTLs undetectable  genomic
prediction
• Look for candidate genes
• markers for selection
• origin of favorable alleles

21. Welcome in Angers, June 22-24 2016
Rosaceae Genomics Conference 8
Thank you for listening