W. Friedt GCIRC General GCIRC General Assembly Assembly , , February February 2009 2009 Manesar Manesar near near Delhi, Delhi, India India Improving Improving rapeseed ( rapeseed ( Brassica Brassica napus napus ) as a highly ) as a highly productive productive o o ilseed and source of valuable products ilseed and source of valuable products for nutrition for nutrition Wolfgang Friedt & Rod Snowdon Plant Breeding Department, IFZ Research Center University of Giessen, Germany
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W. Friedt
GCIRC General GCIRC General AssemblyAssembly, , FebruaryFebruary 2009 2009 ManesarManesar nearnear Delhi, Delhi, IndiaIndia
Improving Improving rapeseed (rapeseed (BrassicaBrassica napusnapus) as a highly ) as a highly productive productive ooilseed and source of valuable products ilseed and source of valuable products for nutritionfor nutrition
Wolfgang Friedt & Rod SnowdonPlant Breeding Department, IFZ Research CenterUniversity of Giessen, Germany
W. Friedt
CONTENT:
Yield potential (Heterosis)
Stability (stress tolerance)
Oil yield and content
Fatty acid composition
Extraction meal quality
Genomics approaches
Outlook & perspectives
W. Friedt
48.0
49.0
50.0
51.0
52.0
53.0
54.0
55.0
56.0
57.0
3.40 3.60 3.80 4.00 4.20 4.40 4.60 4.80
Seed yield (t/ha)
V23-HV24-H
V25-H
MarcantZerucaMaruca
Maplus
Express
Falcon
PantherTalent
Artus
Pronto
ErucicErucic test test hybridshybrids
High erucic cultivars
Standard cultivars (00) Oil
cont
ent(
%)
Seed yield (t/ha)
Oilseed rape: Seed and oil yield
W. Friedt
Investigating heterosis in oilseed rape
Heterosis field trial, Grund Schwalheim, May 2006
Question: Common QTL for heterosis in different oilseed rape crosses?
Material: 2 populations of 250 DH lines and their test hybrids from a common male sterile parent
W. Friedt
Field trials for analysis of yield heterosis
Performance trials in 8 environments (2 years, 4 locations)
Measurement of plant height, plot yield and yield components
Resynthesized (RS) Rapeseed as a Resourcefor Resistance Breeding
Example 1: Clubroot Resistance
- Race specific resistance transferred from B. oleraceainto RS Rapeseed (FU Berlin), by backcrossing
- Simple inheritance (1-2 resistance genes)?
- But, new virulent pathotypes may break resistance
Polygenic types of resistance required
Resistant cultivars for infested locations: Mendel (NPZ)and Tosca (SW Seed)
W. Friedt
- “Complete resistance” against Leptosphaeria maculans/Phomalingam of B. rapa was transferred to RS Rapeseed; cultivars developed by repeated backcrossing and resistance screening
- Monogenic, dominant inheritance- “Breakage” of resistance by virulent pathotypes observed in
Australia
Resistant Spring rape cv. ‘Surpass 400’
Example 2: Phoma Resistance
Resynthesized (RS) Rapeseed as a Resourcefor Resistance Breeding
W. Friedt
Resynthesized (RS) Rapeseed as a Resourcefor Resistance Breeding
Example 3: Novel Verticillium Resistance
Goal: Development of durable Verticillium resistance by combining different sources of resistance; marker-assisted pyramiding of resistance genes
W. Friedt
Screening of mapping parents
0.00
0.50
1.00
1.50
2.00
2.50S
228.
8.1
FS B
1/3.
3S
108.
1.1
K 19
9.16
.2
Sollu
x
R 53
FS 9
4.3
Oas
eSa
mou
rai 1
1.4
Lase
rS
45.2
.2
Gao
you
Expr
ess
YE1
schw
arz
DH-2
6-96
Man
shol
ts 5
.1
Falcon
Expr
ess
Falcon
Expr
ess
617
Lion
YE2
gelb
101
2-98
YE1
gelb
T-2
5629 V8
AU
DP
Cco
rr
Selected RS lines (B.oleracea x B.rapa)B. napus Check cultivars
B. napus Mapping parents
Segregation for Verticillium resistance in Express 617 x R53
‘R53’ (U Göttingen): B. oleraceaacephela (Winter cabbage) x B. rapa pekinensis (Chinese cabbage
Express 617
W. FriedtExpress x R53: 275 DHs, 191 loci (Map from Radoev et al. 2008)
QTL Analysis of Verticillium Reaction
AUDPC
*E32M48b0
*Na10G08b20
*Ol10B0231
*CB1061148
*E32M49u59*CB1006566
*CB1043576
*CB1060987
*MR95112*E32M51k117*E32M51o124*E32M49k129
N15
LOD=7.61Part. R2=29.6
Major QTL explainsapprox. 30 % of resistance phenotype
- DH population Express 617 x R53 (100 DH lines)
W. Friedt
307-406-1 x 307-230-2 Express 617 x R53
BRMS030
Ra2F11_b
AUDPC
AUDPC
N15
Comparative SSR marker analysis
LOD=10.06Part. R2=12.5
LOD=7.61Part. R2=29.6
Major Major resistanceresistance genegene fromfrom B. B. oleraceaoleracea
W. Friedt
Fine mapping of major QTL and marker developmentin Express 617 x R53 via Bulked-Segregant Analysis
Genetic modification of the tocopherolbiosynthetic pathway
Methyltransferase II
2-Methyl-6- phytylchinone
γ - Tocopherol
α - Tocopherol
2,3-Dimethyl-6- phytylchinone
Methyltransferase I
Cyclase
Shikimate Isopentenyl PP
HomogentisateDioxygenase
Phytyl PP
4- Hydroxyphenylpyruvate Geranylgeranyl PP
Reductase
Prenyltransferase
* *
*
* genes have been identified
cyclase
Arabidopsis thaliana
Zea mays
cyclase
HPPD2
Eucaryoticorganism
prenyl
Synechocystis sp.
HPPD1
Arabidopsis thaliana
W. Friedt
Cuphea lanceolata
http://www.mpiz-koeln.mpg.de
Brassica napus
LB p35S t35S pNap tNap pNap tNap RB
npt II clKAS IIIb mut clFATB3
npt II clKAS IIIb mut chFATB2
LB p35S t35S pNap tNap pNap tNap RB
npt II clKAS IIIb mut chFATB2
LB p35S t35S pNap tNap pNap tNap RB
LB p35S t35S pNap tNap RB
npt II clKAS IIIb wt
LB p35S t35S pNap tNap RB
npt II clKAS IIIb mut
T0-Seedling
T1-Plant
T2-Seeds
T2-Plant
T3-Seeds
T3-Plant
T4-Seeds
DevelopmentDevelopment of of transgenictransgenic rapeseedrapeseed plantsplantsand and propagationpropagation of of selectedselected progenyprogeny
Transfer of Transfer of selectedselected genes genes involvedinvolved in in seedseedlipidlipid and and tocopheroltocopherol biosynthesisbiosynthesis to to B. B. napusnapus
((NAPUSNAPUS 20002000 Project)Project)
W. Friedt
name R1 R2 R3 Rel. biolog. activity (%)
α-Tocopherol -CH3 -CH3 -CH3 100
γ-Tocopherol -H -CH3 -CH3 10
δ-Tocopherol -H -H -CH3 3
β-Tocopherol -CH3 -H -CH3 50
TocopherolTocopherol -- StructureStructure
R 3
R 2
OHR 1
CH3
CH3
O
CH3 CH3CH32 4 8
1
345
6
8
7
NAPUS 2000
W. Friedt
HPT= Nap_HPTTC= Nap_TCDouble= DC3_HPPD2,
Nap_HPTTriple= DC3_HPPD2,
Nap_HPT,Nap_TC
N = Number of T2 plants
* P <0,001 (Tamhane)
43344635344235374148401736109N =
1336.557
1336.735
1336.702
1336.699
1336.705
1335.696
1332.515
1332.451
1332.445
1334.691
1334.684
1333.677
1320.642
Kontrolle
1600
1400
1200
1000
800
600
400
200
0
402
Toco
pher
ol(m
g/kg
Öl)
T1-Nachkommenschaft
* * * * * * * * * * * * *100 = 770 ppm
HPT ---TC---- --Double- -------- Triple --------
177189
Variation of Variation of TocopherolTocopherol content in genetically content in genetically engineered engineered seedoilseedoil (pooled T3(pooled T3--seeds)seeds)
Raclaru M. et al. (2006) Molecular Breeding 18:93-107
W. Friedt
CONTENT:
Yield potential (Heterosis)
Stability (stress tolerance)
Oil content and quality
Extraction meal quality
Genomics approaches
Outlook & perspectives
W. Friedt
Influence of Influence of phenolicphenolic compounds compounds of rapeseed mealof rapeseed meal
These compounds contribute to
Dark colourBitter tasteAstringent flavourNon-specific reactions with proteins, enzymes or essential forms that create nutritionally-unavailable products
W. Friedt
GABI-Project: Functional Genomics approaches for the development of yellow-seeded, low sinapine(„YelLowSin“) oilseed rape/canola (B. napus)
Major seed quality QTLMajor seed quality QTLMajor seed quality QTL
Seed colourOil contentProtein contentCellulose
NDF
ADFADL
QTL for seed traits
Verticillium QTL!
W. Friedt
- Use of Arabidopsis-Brassica comparative genomics tools to identify At-regions with synteny to major seed colour QTL
- Isolation, sequencing, marker development and mapping of B. napus orthologs for potential syntenic genes
- Marker saturation in QTL regions of rapeseed
Gene identification: Gene identification: Positional and functional characterization of new seed colour (transparent testa) candidates by At-Bn synteny
Open question: Negative effect of alien germplasm on seed and oil yield in oilseed rape?
W. Friedt
Yellow Rapeseed: OutlookYellow Rapeseed: Outlook
- RT-PCR of CCR-like, CAD-like, CESA9 and TT10 candidates during seed development in black and yellow RILs
- Field trials: New segregating materials for genetic/ marker analysis
- GC-MS to identify major phenolic compound(s) contributing to differences in YE2-DH population
W. Friedt
Finding regulatory genes involved in heterosis:
Many QTL hotspots control heterosis for multiple traits
Epistatic interactions appear to play a role in regulation of QTL
eQTL analysis to identify regulatory candidate genes (e.g. transcription factors) with a key role in heterotic gene expression
Express, V8F1 (Express x V8)
mRNA from seedlings(controlled conditions,
14 & 28 days after sowing)
Selected DH lines withhigh and low heterosis for
cotyledon length and rapeseed yield
Ultradeepexpression
next-gen
EST-Tag profiling
(Illumina/Solexasequencing)
eQTL analysisterosis-relevant
gene expressionof he
W. Friedt
Crop Harvested area Average yield (t/ha) Rate
ha (x106) % arable 1961-1970 1991-2000 dt/year
Cereals 37.8 51 2.6 5.27 0.88
Wheat 18 24 2.4 5.54 1.02
Maize 4.2 6 3.19 8.32 1.69
Rapeseed 3.0 4 1.92 2.88 0.34
Sunflower 1.9 3 1.17 1.54 0.18
Ewert F. et al. (2005) Agriculture, Ecosystems & Environment 107:101-116
Land Land useuse and and selectedselected yieldyield statisticsstatistics forfor majormajorEuropean European graingrain cropscrops
W. FriedtPhoto: Norddeutsche Pflanzenzucht Hans-Georg Lembke KG
AcknowledgementsAcknowledgementsCanadian PartnersCanadian PartnersAAFC & PBI, SaskatoonAAFC & PBI, SaskatoonU of Alberta, EdmontonU of Alberta, EdmontonGenome CanadaGenome Canada
IPB Leibniz Inst, HalleIPB Leibniz Inst, Halle
University of GiessenUniversity of GiessenBenjamin Benjamin WittkopWittkopWilfriedWilfried LLüühshsPanjisaktiPanjisakti BasunandaBasunandaMalteMalte LuhLuh, Stavros , Stavros TzigosTzigosMechthildMechthild SchwarteSchwarte, , a.oa.o..
University of University of GGööttingenttingenHeikoHeiko BeckerBeckerMladenMladen RadoevRadoevWolfgang Wolfgang EckeEcke