Applying Spontaneum

Ahmed JahoorNordic Seed

Denmark

Applying wild barley (H. Spontaneum) in Breeding

The gene pool concept used by plant breeders

TertiaryOther wild barley

Gene pools of Hordeum

SecondaryH. bulbosum

PrimaryCultivated barleyH. spontaneum

Diversity indices within Hordeum vulgare,H. spontaneum and other wild barley species(based on 32 RFLP markers and 223 lines )

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0.70

0.80

0.90

mea

n di

vers

ity (

H)

vulgare spontaneum others

Wild barley in natural habitat

Important traits for breeding from wild barley

• Resistance

• Quality

• yield

Screening of H.vulgare ssp. spontaneum against European and Israeli powdery mildew isolates

0%

10%

20%

30%

40%

50%

60%

70%

80%

R-R R-I I-I R-S S-S

EuropeanIsraeli

Identified and localised new powdery mildew resistance genes with molecular markers in 41 H. vulgare ssp. Spontaneum derived lines

Locus Chromosome Number of alleles

Mla 1H 17

Mli 1H 1

Mlf 7H 7

Mlt 7H 3

Mlj 5H 1

Mlp ? 3

Localisation of qualitative and quantitative inherited mildew resistance gene with their respective number of alleles

italic → localised in H. spontaneum

Distribution of advance lines containing new sources of resistance for winter barley to German breeders

Line Year of distribution

Donor Number of backcrosses

Resistance locus

WI-1 1988 Rs-142-29 4 Mla

WI-2 1988 RS20-2 3 Unknown

WI-3 1989 L74-11 4 Unknown

WI-4 1989 RS2-2 4 Mla

WI-5 1989 RS111-1 4 Unknown

WI-6 1992 RS132-45 4 Not Mla

WI-7 1992 RS122-19 4 Unknown

WI-8 1992 RS170-A-S 4 Unknown

WI-9 1996 RS70-29 4 Mla

WI-10 1996 RS70-30 4 Unknown

Distribution of lines containing new sources of mildew resistance for spring barley to German breeders

Line Year ofdistribution

Donor Number ofbackcrosses

Resistancelocus

SI-1 1987 RS1-12 5 Mla

SI-2 1987 RS81-B-10 5 Unknown

SI-3 1989 1-B-86 5 Mla

SI-4 1989 1-B-87 4 Mlf+mlt

SI-5 1992 RS160-47 5 Linked withMla

SI-6 1992 RS-164-6 5 Mlf+mlt

SI-7 1992 RS14-C-24 4 Mla

SI-8 1996 RS42-8 5 Mli

SI-9 1996 RS42-6 5 Mlt

SI-10 1996 RS137-28 5 Mlf

SI-11 1996 HSY-78 5 Mlj

TILLING procedure

DNA

Isolation

Pooling

Gene specific primer pairs(2 different dyes)

PCR reaction

Denaturation

Renaturation

+ Cel I

Wt Mu Wt Mu

Spectrum of observed fragmentsfor 28 different Mla alleles

D*1B54A Mla16 105 163 222 116 171 343 347 363

RS170-47*Kieb Mla17 105 163 222 91 171 237 313 326 343 347 358 374 415

RS20-1*K.B Mla18 105 163 222 157 171 237 313 343 347 365 374 415

D*1B-86B Mla19 105 163 222 116 171 313 343 347 363

RS145-39*K Mla20 105 163 222 171 313 343 374 398

RS170-10*P Mla25 105 163 222 88 171 300 313 343 374 398

D*1B-20 Mla26 105 222 171 326 343 347 358 365 374 398 400

RS1-8*P.E Mla27 105 222 171 313 326 343 365 374 400 415

D*1B-151 Mla28 150 163 222 171 343 347 363 415

110-4*S Mla29 105 163 222 116 171 313 343 347 374 419

Turkey290 Mla31 105 163 222 91 116 171 237 313 343 347 374 415

RS90-13 Mla33 105 222 88 171 300 313 343 347 363 365 398 415

RS70-29xSxCxSxH Mla34 105 163 222 171 313 343 374 415

RS2-2xPicc. Mla35 105 163 222 126 171 313 343 374

P04A Mla7, Ml-k 105 163 222 88 171 300 313 343 347 374 415

Pallas Mla8 105 163 222 171 313 343 347 374 415

P08B Mla9 105 163 222 171 286 313 343 347 374 411

P07 Mla9, Ml-k 105 163 222 88 171 300 313 343 347 374 415

OPTIC Mla12 105 163 222 88 171 300 313 343 347 374 415

D*1B54A Mla16 105 163 222 116 171 343 347 363

RS170-47*Kieb Mla17 105 163 222 91 171 237 313 326 343 347 358 374 415

RS20-1*K.B Mla18 105 163 222 157 171 237 313 343 347 365 374 415

D*1B-86B Mla19 105 163 222 116 171 313 343 347 363

RS145-39*K Mla20 105 163 222 171 313 343 374 398

RS170-10*P Mla25 105 163 222 88 171 300 313 343 374 398

D*1B-20 Mla26 105 222 171 326 343 347 358 365 374 398 400

RS1-8*P.E Mla27 105 222 171 313 326 343 365 374 400 415

D*1B-151 Mla28 150 163 222 171 343 347 363 415

110-4*S Mla29 105 163 222 116 171 313 343 347 374 419

Marker for qualitative traits

std. susc. res. res.susc.

STS marker for barley powderymildew resistance gene Mla12developed from AFLP marker

std. std.res.uncl.

susc.ScrFI

susc.uncl.

res.ScrFI

SCAR marker for barley powderymildew resistance gene Mla12developed from RFLP marker

Released varieties developed with the new sources o f resistance against powdery mildew originating from H.

spontaneumSally Streng 1999 SI-1Roxana Breun 1998 1-B-53Vrena Nordsaat 2000 WI-1Havanna Breun 2000 1-B-53Alabama Lochow-Petkus 2000 SI-8Philadelphia Lochow-Petkus 2001 SI-8Marnie Breun 2002 1-B-80Venezia Bauer 2002 WI-1Denise Breun 2003 1-B-53Antalya Breun 2004 WI-7Djamila Nordsaat 2003 SI-4Grace Dr. J. Ackermann 2008 1-B-53Henrike Nordsaat 2007 1-B-53Isotta Breun 2004 1-B-53Iron Nordic Seed 2008 1-B-53

Leaf rust, powdery mildew and scald resistance genes on barley chromosome 2H 20 cM

S LC

[Rph1] MlLa

qLr1 qLr2 qLr3 qRh1

qPm3

Name

Population

LODEffectres. ParentVar.expl.

Krona xHOR106310.92.13 UnitsHOR106311%

qLr1

Krona xHOR10634.41.6 UnitsHOR106324%

qLr2

Krona xHOR10633.50.5 UnitsHOR10632%

qLr3

Igri xDanilo16.91.1 pointsIgri32%

qRh1

1B-87 xVada3.814 UnitsVada6%

qPm3

Leaf rust, powdery mildew and scald resistance genes on barley chromosome 5H

S C LRph2 RphQ Rph9 Rph12

20 cM

Name

Population

LODEffectres. ParentVar.expl.

1B-87 xVada5.919Vada9%

qLr6

1B-87 xVada15.952Vada41%

qLr5

1B-87 xVada6.517 unitsVada7%

qPm6

qLr5 qPm7

qPm6qLr6

qLr7

1B-87 xVada7.718 units1B-879%

qPm7

1B-87 xVada7.0151B-8741%

qLr7

Mlj

QTL analysis of powdery mildew resistance in a cross between wild barley and cultivated barley

m32p31-040m59e41-0410m48e38-2315MWG36MWG219720m31p31-0826m47e38-1336m32p31-1744MWG7549MWG80064MWG5567m59e41-0671S-921773S-11275BMag211m59e36-1277m47e41-0279m47e38-1285m47e38-1987BMac13101MWG943106m47e32-19112MWG518118BMag382127cMWG695132cMWG706a135m48e32-08137m48e38-20149

m47e41-14171m41p31-12174MWG2028177m33p31-11182MWG912188m31p31-07189m47e38-14190

1H

S-T650S-133-23S-2366cMWG68211m41p31-0215BMac13423

m48e32-0336m47e32-0643

m32p31-1153

MWG6463m42p31-0669

2H

m31p31-250S-92046m31p31-1811MWG6519m41p31-1822m47e38-1026m59e38-3127m48e32-1528m47e38-1533cMWG70236m48e41-0343HVBKASI49m48e32-0152m48e32-1354BMag37864m41p31-0881MWG503a87m33p31-1889m41p31-0792S-12098m59e36-08108m59e36-07116S-217118m31p31-17124MWG82126MWG46130

m59e36-180m32p31-033cMWG6914MWG63b10m33p31-0518m59e38-0833m59e36-0434S-reg1937MWG58439

m48e38-1558m48e38-0159m47e32-2062m59e38-2964cMWG68070m48e38-1674m47e32-1880BMag22592MWG555b100

m41p31-14119

MWG961133BMag13147m59e41-15153m59e41-16154S-L8156m41p31-11159m31p31-15168m41p31-09170m59e38-26183

m41p31-01193

3Hm31p31-060m48e38-115m59e38-0112m59e36-0113m47e38-0318m33p31-0422m33p31-1523MWG7724MWG203329WHS17841m32p31-1653MWG5857BMac18163m48e38-1270m47e38-1176m48e38-0779m47e32-0285m48e32-2187m48e32-0590m48e32-1092m47e41-0499MWG57103m47e41-10110m48e32-17128

m48e38-18144m47e38-23146WMS6150m33p31-12159HVM67162

4Hm32p31-010MWG5302BMag2066m48e32-1110MWG56420m42p31-0722m33p31-1424MWG4a36MWG1843MWG83651m59e36-0962MWG3965m42p31-0375MWG62676m48e38-0679m47e32-0881BMag32182MWG649a83m48e32-2285m47e41-0788m47e32-0499m31p31-24107MWG53109m41p31-20114m42p31-01121cMWG633127MWG539135m32p31-06136m41p31-19141S-9203151

7H

qml1qml2

qml3

qml4qml5

P < 0.0005

Mla

MlLa

Mlf

Leaf rust, powdery mildew and scald resistance genes on the barley chromosomes

1H 2H 3H 4H 6H 7H

RhyRh

qRh2

qRh3

Rrs13

qRh4Rh2

qRh8

5H

Rph4

[Rph1]

qLr2

qLr1

qLr3

[Rph7]

[Rph11]

qLr4

qLr9

Rph2RphQ

Rph9Rph12

qLr5qLr6

qLr7

MlaMlk

Mlnn

MlGa

qPm1qPm2

MlLa

qPm3

Mlg

mlo

qPm4

qPm5

MlhqPm8

qPm9

mlt

Mlf

qPm10

qPm11

qPm12

qPm6qPm7

Mlj

Malting qualty traits detected in wild barley

Trait a

QT

L in S42IL

Chr. b

Introgression (in cM) c

Bin range

d

Introgression line

Effect e

LSM

EA

NS

(IL) f

Diff. g

RP

[S42IL] (%

) h

QT

L in parent population S

42i

Candidate genes

j

AA

QAa.S42IL-4H.a 4H 14–31 2–4 S42IL-116 L + I 546.9 91.3 20 QAa.S42-4H.a

QAa.S42IL-4H.b 4H 170–190 12–13 S42IL-124 L + I 529.4 73.9 16 .2 QAa.S42-4H.c Bmy1 1

GSF

QGsf.S42IL-2H.a 2H 17–42 2–4 S42IL-107 L + I 86.4 6.7 8.4 Ppd-H1 6

QGsf.S42IL-2H.c 2H 80–86 7–8 S42IL-110 L 85 5.3 6.7

QGsf.S42IL-4H.a 4H 31–95 4–8 S42IL-119 L 84.6 4.9 6.2

KOL QKol.S42IL-4H.a 4H 14–31 2–4 S42IL-116 L 46.4 2.2 5.1

Schmalenbach and Pillen, 2009

QTL detected for agronomic traits in wild barley

TraitaQTL in S42IL

Chr.bIntrogression (in cM)c

Introgression line

EffectdLSMEANS (IL)e Diff.f RP (IL) %g

EARQEar.S42IL-6H.a 6H 145–155 S42IL-132 I 1359.0D07 391 40.4

HEAQHea.S42IL-1H.a 1H 0–85 S42IL-102 L + I 68.2 1.9 2.9

HEIQHei.S42IL-1H.a 1H 70–85 S42IL-105 L + I 89.9 9.1 11.3

LAHQLah.S42IL-1H.a 1H 70–85 S42IL-105 I 6.3G07 3.2 100

YLDQYld.S42IL-4H.a 4H 31–57 S42IL-118 I 73.5D07 7.8 11.8

Schmalenbach et al. 2008

No. Favorable 1) Max. RP 2) Difference Trait QTLs exot. alleles [Hsp:Hv] [Hsp]–[Hv]

Heading 10 8 - 7.9 % - 5.7 d

Yield 12 3 + 7.1 % + 4.0 dt/ha

Powdery 8 6 - 51.5 % 2.5 scoremildew

1) BC2DH lines with the exotic genotype [Hsp] are on a verage superior to lines with the elite barley genotype [Hv] at a given marker locus.

2) Performance of homozygous exotic genotypes relative to homozygous elite barley genotypes at a marker locus.

Localization of QTL regions and favorableexotic QTL alleles in S42

Pillen et al.

Summary of QTL verification with 39 S42-ILs

- Hsp allele reduces trait+ Hsp allele increases trait

= QTL verifiedPillen et al.

0

1420

28

39

5263

6870

75

85

105115

130

1H

144

65

162

30

25

49

65

94

100

110

130

155

165

175

190

3H

70

142125

31

44

57

80

95

125

130131132

150

170

180

190

4H

55

17

27

92

107

122

139

143

146

159

2H

42

8680

67

0-14

14-3

9

39-7

070

-85

52-7

0

17-9

2

17-4

2

17-2

767

-92

80-8

6

65-7

0

155 -

190

155 -

175

155 -

165

100 -

130

14-4

4

31-5

7

80-9

5

31-9

5

14-2

517

0 -19

0

125 -

132

125 -

170

55-5

7

20

67

190

4480

95

5H0

12

24

43

48

69

85

126

137

162

165

43-4

816

2 -16

569

-85

69

6

40

96

103

107

112

135

145

155

6H

40-1

12

96-1

12

112 -

155

135 -

155

145 -

155

640

9611

2

19

27

50

62

75

9394

100

120

133

146152155

166

178180181

7H

5062

-75

75-1

55

146 -

166

146 -

155

166 -

181

178 -

181

133

Regions of chromosomes so far without ILsFinished IL Finished Sub- IL

(Schmalenbach et al. , unpublished)Days to heading

*) favorable exotic allele

HEA

ILs selected for heading date QTLs in GABI-Malt

HEA*

HEA*

HEA

HEA*

HEA*

HEA*

Conclusion

• Wild barley has been shown to posses new and effective sources of disease resistance

• Transfer of resistance gene from wild barley to cultivated barley has been achieved successfully

• QTL for yield and malting quality have been detected in wild barley.

• Future will show whether these QTLs could be transferred successfully in the cultivars

Acknoledgment

• TUM- Weihenstephan

• Risø National Laboratory (Nina M. Jensen)

• University of Copenhagen (Gunter Backes)