Development of molecular markers for application in A stralian Canola application in Australian Canola

breedingg

C l M l l M k PCanola Molecular Marker Program(harsh.raman@industry.nsw.gov.au)

Mapped Blackleg Resistance lociMapped Blackleg Resistance lociSkipton/Ag Spectrum (191 lines) Skipton/Ag-Spectrum (191 lines) BLN2762/Surpass400 (139 lines)

C l b *3/W t 10 (76 li ) Columbus*3/Westar 10 (76 lines)

Maxol*1/Westar 10 (101 lines)

Ag-Castle/Topas Ag-Castle*4/Westar10 (173 lines) g ( )

Mapped blackleg resistance loci in the DH population from BLN2762/Surpass-400population from BLN2762/Surpass-400

Screened with three isolatesScreened with three isolatesD1 (AvrLM2,5,6,9)D7 (AvrLM1 3 5 6 8)

r2=70%D7 (AvrLM1,3,5,6,8)D10 (AvrLM5,6,8,9,S)

Rlm9M1

M2

A10A7

Mapping of Ag-Castle/Topas

Population is developed by International

Mapping of Ag Castle/Topas

Population is developed by International Blackleg Consortium, Canada

F -1 F1-1 F1-2 Mapped with SSR (Canada Wagga) and Mapped with SSR (Canada, Wagga) and

DArT markers (Wagga) Qualitative Resistance Genes Qualitative Resistance Genes Quantitative Resistance Genes

3-4 QTLs– 3-4 QTLs

Strong Evidence of novel genesStrong Evidence of novel genes

QTL on chromosome X: r2= 28% – Field conditions

QTL on chromosome X: r2= 25%– Seedling stage

Validation of Loci for BLR

Bi-parental populations– Evaluate two alleles/locus

•Skipton/Ag-Spectrum•Maxol/WestarA C l /• Known ancestry •Ag-Castle/Westar

•Surpass/Topas•BLN3347/Carousel-10

Association mapping– Evaluate more alleles/locus

• Historic recombination

Digital Image analysis

BR Scores of 188 Lines Using Differential SetBR Scores of 188 Lines Using Differential SetD1 D2 D3 D4 D5 D6 D7 D8 D9 D10

4.730140304 7.646925 7.128249 6.971014 1.836446 2.285203 2.115384 3.319144 4.757586 6.789557

7.097330341 6.618246 7.920466 6.133153 4.431464 6.812594 5.135307 3.122412 3.744606 7.372861

4.356579722 7.232666 7.533186 6.021005 4.989769 4.887729 5.961428 3.576325 3.821709 6.558984

3.668504598 6.394026 6.921822 6.13768 3.952241 5.087057 4.581889 3.161766 4.912204 6.454949

4.522183939 6.229967 6.028761 6.361742 2.957218 1.091836 2.472812 1.323677 5.722262 4.500186

6.015414023 6.040849 6.505169 6.073434 2.411565 2.724239 4.632233 3.208622 3.200387 5.417817

5.591654768 7.413355 7.361577 6.63768 3.654039 2.698503 2.850807 2.183013 5.962158 6.839883

4.423611703 6.555453 7.396042 5.633161 1.550735 3.589092 5.679019 3.309156 4.056738 6.270415

6.886679493 7.023896 7.984523 6.358386 5.292574 6.127917 4.005167 4.99292 6.186199 7.152023

6.135385064 7.462896 7.961975 6.679347 4.994903 6.960856 4.823716 4.126742 5.141677 7.428624

4.129040146 5.409193 6.290693 4.01268 2.825122 3.857382 1.85182 2.110125 5.153192 6.457673

5.431069152 5.915106 4.398745 1.204228 1.863305 4.074617 4.304964 2.816818 3.277779 4.295161

5.614302808 6.066947 7.552231 5.31672 3.201049 6.393255 4.565232 3.339089 5.958782 5.880304

2.826531339 6.369425 6.558874 4.349931 3.370806 3.242903 1.600677 3.854578 4.823295 3.60767

4.073260062 6.035585 7.189969 4.833161 3.093878 4.880359 5.526491 3.301663 2.586679 6.747776

3.520257892 6.004412 5.034623 6.961948 2.380899 5.700361 1.953367 1.297054 3.235428 5.006055

2.362642866 5.175833 6.251495 3.34907 3.678294 3.312643 3.117591 3.749729 3.562106 3.181641

Evaluated 188 lines for Adult Plant Resistance

AV-Garnet (Rlm1 9) AV Garnet (Rlm1,9) Monola76 (RlmS) ATR-Cobbler (Rlm 4,9)( , ) CB-JardeeHT (Rlm2,3) Hyola50 (Unknown)

Results: S B j d B Some B. juncea and B. carinata lines were resistant to all stubbles

Phenotypic Relationships for BLRGroup average

Resemblance: D1 Euclidean distance

100

ance

50

Dis

ta

0

Hyo

la50

Hyo

la76

AT

C931

84-1

AT

C940

44-1

CB

J-00

1S

eeth

aA

TR-S

igna

lA

v-Ru

byT

orna

doT

T45

C05

Bra

voT

TO

asis

CL

ATR

-Bar

raA

v-Ja

deT

awrif

ficT

TAT

R-S

umm

ittSA

RDI

607

Ag-C

omet

Ag-

Spec

trum

Car

ouse

l-10

Tar

cool

a46

C04

Sto

rmT

TS

urpa

ss40

2CS

urpa

ss40

046

Y78

ZY0

10B

LN27

37*C

O02

0305

-P71

-11

P624

Wes

bell

RO

Y33-

99P

3Ag

-Mus

ter

BLN

3347

CB-T

elfe

rR

OY

9712

5B

LN28

52-0

3W02

RO

Y33-

99P

2Iw

ao-n

atan

eA

v-O

pal

Hur

rican

eTT

Nor

in_22

RO

Y33-

99P

4B

LN33

03*C

R030

2B

LN36

14R

OY3

1-99

P1

Thu

nder

TT

Osc

arR

OY

9705

6SA

RDI

604

Lant

ern

Rot

tnes

tTT

CC

B_T

rilogy

Rive

tte

AT

R409

BLN

3343

*CO

0401

CB-A

rgyl

eSu

rpas

s501

TT

44C

73D

unke

ld45

C75

Mys

ticA

TR-

Cob

bler

Rock

etC

L45

Y77

AT

R-B

eaco

n44

Y06

War

riorC

LF

linde

rsT

TC

Hyo

la60

ATR

-Mar

linP6

1703

-p74

-4A

G-O

utba

ckW

A05

0088

WA

0500

89Ex

pand

erR

ainb

ow69

1198

04-p

34Su

rpas

s603

CL

ZY0

15A

v-S

apph

ireC

harlt

onW

A05

0097

ZY0

0503

-p74

-6C

B-B

oom

erZ

Y016

RO

Y98

279

03-p

74-3

Fan

023

Yu17

8R

OY

9700

3Ch

on-n

amZ

hong

shu-

angN

0.4(

3087

2)C

olt

Lisor

a03

-p74

-11

ZY0

13A

-198

90Z

Y001

RO

Y97

019

ZY0

14A

TR-

Stu

bby

GAS

GO

TTT

ERI

(OO

)R9

06-P

71-1

Dra

kkar

GSC

-5P3

083

RO

Y33-

99P

5D

ing47

4T

atyo

onQ

u110

4W

esre

o06

-P71

-2M

arno

oZ

Y003

ZY0

08Z

Y009

RO

Y33-

99P

1R

OY

9701

7M

aluka

ZY0

12F

an16

8N

indoo

BLN

3343

*CO

0402

46C

76C

B_P

ilbar

aA

TR-B

anjo

Skip

ton

RO

Y97

033

Scoo

pY

icka

dee

Zho

ngsh

u-an

gN0.

4(30

920)

RO

Y97

018

RO

Y97

021

Wes

bark

erR

OY3

3-99

P6

Wes

roon

aW

A05

0096

Eur

eka

RS

O94

-67(

98R

OY3

3-99

P8

Gro

use

ZY0

04A

TR-H

yden

BLN

1990

Zho

ngyo

u-za

ngN

o.8

Zhon

gyou

_821

Ding

110

CB-

Tan

ami

Fan

189

Bar

ossa

6711

09H

yola

61M

ajor

Mut

u-98

-1Z

Y002

Wes

way

95*3

06-3

10R

OY

9709

6Ce

scalj

arni

Mon

tyS

hiral

eeA

G-E

mbl

emT

ranb

yR

OY

9833

0Ra

nge

05-P

36_R

RO

Y97

020

CB-T

rigol

dR

OY

9707

3Z

Y007

N00

1-28

-246

Karo

oR

OY3

3-99

P7

Samples

0

Principal Component Analysis Principal Component Analysis 50

05-P71-1146Y78

BLN1990 P624

ZY010

0671109

05-P36_R

06-P71-1

44C73

44Y06

45C75

45Y77

46C04

46C76

95*306 310

AG-Emblem

Ag-Muster

ATR409

ATR-Banjo

ATR-Barra

ATR-Beacon

ATR-Cobbler

ATR-Hyden

ATR-Marlin

ATR-Stubby

ATR-Summitt

Av-Opal

Barossa

BLN1990

BLN2737*CO0203

BLN2852-03W02

BLN3303*CR0302

BLN3343*CO0401

BLN3343*CO0402

BLN3614

Carousel-10

CB-Argyle

CB_Pilbara

CB-Tanami

CB_Trilogy

Cescaljarni

Ding110

Ding474

Drakkar

Dunkeld

Eureka

ExpanderFan168

Fan189

FlindersTTC

Grouse

GSC-5

Hyola50

Hyola60

Hyola61Hyola76

Karoo

Lantern

Monty

Mystic

N001-28-246

Norin_22

Oscar

P3083 Rainbow

Range

RivetteRocketCL

RottnestTTC

ROY31-99P1

ROY33-99P1

ROY33-99P4

ROY33-99P5

ROY33-99P6

ROY33-99P7

ROY33-99P8

ROY97017

ROY97018ROY97020

ROY97021

ROY97056

ROY97073

ROY97125

ROY98330

RSO94-67(98SARDI604

SARDI607

Shiralee

StormTT

Surpass501TT

Surpass603CL

Tarcoola

Tatyoon

TawrifficTT

ThunderTTTranby

WA050096

WarriorCL

Wesbarker

Wesbell

Wesroona

WeswayYickadee

Zhongshu-angN0.4(30920)

Zhongyou_821

Zhongyou-zangNo.8

ZY004

ZY007

0.5%

)PC

2

69119803-p74-11

03-p74-3

03-p74-4

03-p74-6

04-p34

06-P71-2

45C05

46C0495*306-310

A-19890 Ag-Comet

AG-Outback

Ag-Spectrum ATC93184-1

ATC94044-1

Av-JadeAv-Ruby

Av-Sapphire

BLN3347

BravoTTCB-Boomer

CB-TelferCB-Trigold

CBJ-001Charlton

Chon-namColt

Expander

Fan023GASGOTT HurricaneTTIwao-natane

Lisora

Major Maluka

Marnoo

Mutu-98-1Nindoo

OasisCL

P617Qu1104

ROY33-99P2

ROY33-99P3ROY97003

ROY97017ROY97019ROY97033

ROY97096

ROY98279

Scoop

Seetha

Skipton

Surpass400

Surpass603CLTERI(OO)R9

TornadoTT

WA050088WA050089

WA050097

WesreoYu178

Zhongshu-angN0.4(30872)

ZY001

ZY002

ZY003

ZY005ZY008

ZY009

ZY012

ZY013ZY014

ZY016

(10

-50

ATR-SignalSurpass400

Surpass402C

ZY015

80

90

-100 30

40

50

60

70

80

Eig

enva

lue

%

-100 -50 0 50 100 150PC1(67.6)

1 1.5 2 2.5 3 3.5 4 4.5 5 5.5Component

0

10

20

Non-metric Multi-Dimensional ScalingNon metric Multi Dimensional ScalingResemblance: D1 Euclidean distance

DistanceSurpass402C

2D Stress: 0.11

60Surpass402C

03-p74-6

06-P71-2

44C73

45C05

46C76ATC93184-1

ATR409 ATR-Banjo

ATR-Barra ATR-Beacon

BLN3343*CO0401BLN3343*CO0402

BravoTT

Carousel-10CB_PilbaraCB_Trilogy CharltonDunkeld Fan168

L t MarnooOasisCL

Oscar

Qu1104RivetteRottnestTTC

ROY33-99P1ROY97017

ROY97033ROY97056

SARDI604

Scoop

Skipton

Tarcoola

TawrifficTT

Wesreo

ZY005 ZY008ZY009

ZY012

ZY015

67110903-p74-11 03-p74-3

03-p74-4

05-P36_R06-P71-1

44C73

44Y06

45C75

45Y77

46C04

95*306-310 A-19890Ag-Comet

AG-Emblem

Ag-Muster

AG-OutbackAg-SpectrumATC94044-1 ATR-CobblerATR-HydenATR-Signal

ATR-StubbyATR-Summitt

Av-Jade

Av-Opal

Av-Ruby

Av-Sapphire

Barossa

BLN1990

BLN2852-03W02

BLN3303*CR0302

BLN3347

BLN3614

CB-Argyle

CB-Boomer

CB-Tanami

CB-TelferCB-Trigold

CBJ-001

Cescaljarni

Chon-namColt

Ding110

Ding474

DrakkarEureka

Expander

Fan023

Fan189

GASGOTT

Grouse

GSC-5HurricaneTT

Hyola61

Iwao-natane

Karoo

Lantern

LisoraMajor

Maluka

Marnoo

Monty

Mutu-98-1

Mystic

N001-28-246

NindooNorin_22

P3083

P617

RainbowRangeRocketCL

ROY31-99P1

ROY33-99P2

ROY33-99P3

ROY33-99P4

ROY33-99P5

ROY33-99P6ROY33-99P7

ROY33-99P8

ROY97003ROY97018ROY97019

ROY97020ROY97021

ROY97073

ROY97096

ROY97125

ROY98279

ROY98330

RSO94-67(98

SARDI607

Seetha

Shiralee

StormTT

Surpass501TT

Tatyoon

TERI(OO)R9

ThunderTTTornadoTT

Tranby

WA050088

WA050089

WA050096

WA050097

WarriorCLWesbarker

WesroonaWesway

Yickadee

Yu178

Zhongshu-angN0.4(30872)

Zhongshu-angN0.4(30920)

Zhongyou-zangNo.8

ZY001

ZY002

ZY003

ZY004

ZY007

ZY008ZY012

ZY013

ZY014ZY016

691198

04-p34

05-P71-11

46Y78

ATR-Marlin

BLN2737*CO0203FlindersTTC

Hyola50

Hyola60

Hyola76

P617P624

Surpass400

Surpass603CLWesbell Zhongyou_821

ZY010

Characterisation of APR linesCharacterisation of APR lines

Differential set was used to Differential set was used to characterise APR linesAll li i t t t ll All lines were resistant to all isolates

Loci remains to be characterised

Conclusions

Mapped and validated known Rlm d th l i i t d ithand other loci associated with

blackleg resistanceCharacterised germplasm for

blackleg resistance at seedling andblackleg resistance at seedling and adult plant stages– Identified new sources and loci for

blackleg resistance

AcknowledgmentsgCanola Molecular Marker Program

Collaborators •Australian Canola

I&I NSW•Rosy Raman

Marker Program•Simon Diffey (NSWDPI)•Steve Marcroft (MGP)•Kurt Lindbeck (I&I NSW)

Breeding Programs

•International Blackleg Consortium, C d•Rosy Raman

•Belinda Taylor•Ata Rehman

Kurt Lindbeck (I&I NSW)•Angela Van de Wouw (UM)•Paul Eckerman (UA)

Canada

Ata Rehman•Neil Coombes•David Luckett

•Ray Cowley (I&I NSW) •Dave Roberts (I&I NSW)

UQ•Jacqui Batley•David Edwards

•Shanoor Hossain (VicDPI)•Phil Salisbury (UM)B b (Vi DPI)•David Edwards

•Jiri Stiller•Bob (VicDPI)