Discovery of QTLsin apple by using thepedigree-based approach (PBA)

T. Letschka, G. Pagliarani, L. Lozano, W. Guerra, M.C.A.M Bink, J. Kruisselbrink, R. Voorrips, M. Di Guardo, E. Di Pierro, E. Van de Weg

The pedigree based approach (PBA)

full sib familiesparentsgrand parents & founders

GoldenDel

AntonovkaOB

F_B8_34.16

Clochard

Winesap

F_Delicious

F2_26829-2-2

EsopSpitz

F_Jonathan

Prima

Wagenerap

X-6398

ReiDuMans

GranSmith

Coop-17

Cox

RomBeauty

F_O53T136

RallsJan

PRI668-100

RedWinter

McIntosh

JerseyBla

F_Wealthy

F_LadyWill

DrOldenbu

F_Alwa

F_JamesGr

WorcPearm

F_Braeburn

BX44.14

BeautBath

I_M01

I_W01

12_N01

12_P01

12_L01

I_BB02

12_F02

12_J01

I_J01

Z185

PRI612-1

Idared

PRI672-3

X-3259

X-6679

Crandall

KidsOrRed

Fuji

Melrose

Wealthy

PinkLady

Pinova

Alwa

Lambourne

Braeburn

Rewena

Pirol

Reanda

DiPr_001

X-4355

Florina

X-3177

X-6799

O53T136

Gala

X-2771

X-4638

PRI54-12

Sampion

FuPi_001

TeBr_004

RePir_110

AlPi_08

PiRea_083

X-6820

X-6681

Baujade

TN_R10A8

X-6564

Galarina

X-3143

X-3263

X-6808

Dorianne

Liberty

FuGa_002

GaCr_02

GaPi_03

PiGa_01

AlSz_01

BVIII_34.16

F_X-4355

Chantecler

Delicious

PRI14-126

PRI14-152

Jonathan

Wines_Prima

Rubinette

X-6823

Macoun

LadyWill

Alkmene

Clivia

JamesGr

Telamon

67-47

Discovery

X-6683

X-3305

X-6417

X-3318

12_O03

12_K01

I_CC03

12_I02

MeLi_01

RedWinterX3177

YOUR LOGO

Origin of plant material

>1,500 individuals from 7 different countries

>1,200 individuals in 25 families

• phenotyped for 3 consecutive seasons

• at 4 time points each season (HiDRAS)

• >30 different traits from pre- to post-harvest

• genotyped with Illumina 20K SNP array

• data scrutinizing

• consensus map

Genotypic dataset

• 7K SNPs in 1,112 haploblocks through Haplotyper• Marker density: mean interval = 1 cM

Trait under investigation

Titrable acid after 2 months of cold storage and 2 weeks of shelf life.Average of 3 phenotyping seasons.

Already published data

LG 1: Xu et al. 2012 (2 half sib pops) -> M3 geneLG 6: Xu et al. 2012 (2 half sib pops) -> M2 geneLG 8: Liebhard et al. 2003 (Fiesta x Discovery)

Kenis et al. 2008 (Telamon x Braeburn)Kumar et al. 2012 (genomic selection)

LG 16: Maliepaard et al. 1998 (Prima x Fiesta) -> Ma locusLiebhard et al. 2003 (Fiesta x Discovery)Kenis et al. 2008 (Telamon x Braeburn)Xu et al. 2012 (2 half sib pops)Kumar et al. 2012 (genomic selection)Khan et al. 2012 (Malic acid transporter gene Ma1 & Ma2)Bai et al. 2012 (functional SNP marker in Ma1)

Kenis et al. 2008 (Telamon x Braeburn): LG2, LG10, LG13, LG15, LG17

Bai et al. 2012

QTL positions and Bayes factors

~49cMBF:30

~13cMBF:11

~1cMBF:13

~31cMBF:27

~13cMBF:6

~9cMBF:29

~13cMBF:4

~57cMBF:4

QTL effect sizes

Additive effect size of all QTLs sums up to 2.04

=> explains 55% of phenotypic variance

Genotypes amongst full sib parents

Genotypes amongst ancestors and founders

Genetic and physical position of Ma1

Malus x domestica v1.0

Ma2 Ma1

HB-16-2e HB-16-2f

Favourable alleles on LG 1

Favourable alleles on LG 1

RedWinter Jonathan Wagenerap Prima GoldenDel Clochard Coop-17 RallsJan Winesap X-2771

Idared F_X-4355 Chantecler Z185 Florina Delicious X-3143 X-6823

X-3177 X-3259 X-4355 X-6820 Fuji X-6679

X-3263 RedWinterX3177 X-6683 X-3318

12_I26 12_I29 12_I30 12_I32 12_I33 12_I34 12_I35 12_I36 12_I37 12_I38 12_I43

2 4 4 2 2 9 3 2 4 4 5 4 4 3 4 2 7 4 2 3

3 38 83 3

3 38 83 3

2 34 84 3

3 38 83 3

2 34 84 3

2 34 84 3

3 38 83 3

2 34 44 4

2 34 84 3

3 28 43 4

2 34 44 4

2 34 84 3

2 32 82 3

2 32 82 3

2 32 84 3

2 34 84 3

2 34 84 3

2 22 22 4

* ** ** *

2 22 22 4

2 32 84 3

2 24 24 2

2 2* *4 *

2 24 44 4

2 22 24 2

3 28 *3 2

2 22 24 2

2 32 84 3

2 22 44 4

2 22 4

2 24 2

2 22 12

3 28 4

2 22 4

2 22 4

2 34 8

2 24 2

2 22 2

2 32 8

LG 1 – QTL clustering

TA

Brix

Russeting Attractiveness

Firmness

Fruit size

Observation

• QTLs for different traits co-localize on LG 1

• within 5 cM

• immediately downstream of Rvi6 haploblock

Interpretation

• Is it a pleiotropic effect?

• Are the traits correlated with each other?

• Is there a link to Rvi6 resistance? Linkage drag?

Results are highly reliable• thorough scrutinizing of input data (genotyping, mapping, phenotypes...)

• Short linkage groups

• Marker consistency

• Hardly any double recombinants

• Consistency between generations

• No warnings during QTL analysis runs

• Narrow QTL regions that contain known target genes

Conclusion I

Conclusion II

• According to FlexQTL, the phenotypic variance in titrable acid is explained in

a multiple QTL model that involves:

5 decisive QTLs + 1 strong QTL + 2 positive QTLs

• The additive effect of these QTLs accounts for 55% of the observed

phenotypic variance

• Most important QTLs on LG 16, LG 8, LG 1.

• Ma1 & Ma2 could be pinpointed within the QTL region on chromosome 16.

• QTLs for various different quality traits cluster on LG 1, in addition to scab

resistance.

Outlook

• Move closer to Ma1 gene

• Investigate on ‘pleiotropic effect‘ on LG 1

• Analyse other QTLs related to TA and Brix

• Examine different time points regarding post harvest

• Compare results to sensorial traits

• Extrapolate markers to be used for marker-assisted breeding

Acknowledgements

HiDRAS phenotypersgenotyping facilitymappersMario Di Guardo (SNP scoring)Erica Di Pierro (consensus map)Roeland Voorrips (Haplotyper)Hans Jansen (integration of newly phenotyped populations)Johannes Kruisselbrink (Visual FlexQTL)

Giulia PagliaraniHélène Muranty, Stijn VanderzandeWalter Guerra

Marco BinkEric van de Weg

Thank you for your kind attention.