mapping and detection of downy mildew and botrytis bunch...
TRANSCRIPT
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Chin-Feng Hwang, Ph.D. State Fruit Experiment Station
Darr College of Agriculture
Mapping and Detection of Downy
Mildew and Botrytis bunch rot
Resistance Loci in Norton-based
Population
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Norton has been grown in
Missouri for over 160 years,
but little is known about the
genetics of its disease
resistance, cold hardiness
and berry quality.
Vitis aestivalis-derived
‘Norton’
State Grape of Missouri
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Norton has naturally evolved resistance to
1. Powdery mildew
2. Downy mildew
3. Berry rot complex including Botrytis
Bunch rot, Bitter rot and Black rot
4. Insect Phylloxera
5. Cold hardiness
Genetics of Norton (Missouri State) Grapes
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Vitis aestivalis-derived ‘Norton’ Cold hardy and Resistant to most fungal pathogens
Good wine quality
Vitis vinifera ‘Cabernet Sauvignon’ Cold sensitive and Susceptible to most fungal pathogens
Great wine quality
A need exits to breed for grapevines that
would combine the superior wine quality
of V. vinifera with the disease resistance
and cold hardiness of Norton.
Norton vs Cabernet Sauvignon
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Interspecific Hybrid Identification
Genetic profiles (allele sizes in bp) of grape varieties at various SSR loci
Scientia Horticulturae (2014) 179: 363
Crosses # Plants evaluated
# True hybrids
% True hybrids
Norton (♀) x Cabernet Sauvignon (♂) 286 252 88.1
Norton (♂) x Cabernet Sauvignon (♀) 24 21 87.5
Crosses tested for interspecific hybrid production
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Genotyping
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Norton Linkage Map Construction
1. 1,157 SSR markers were test on the parents
and 4 progenies for polymorphism
2. 414 polymorphic markers were identified and
screened through a-182 genotype
population
3. 411 markers clustered in 19 linkage groups
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1 2 3 4 5 6 7 8 9
10 11 12 13 14 15 16 17 18 19
Molecular Breeding (2017) 37:64
There are 26 gaps larger than 10 cM.
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VitisGen I Project (2011-2016)
• A 5-year project funded by the USDA-National Institute of
Food and Agriculture (NIFA) Specialty Crops Research
Initiative
• Combines the expertise of breeders, geneticists,
pathologists, physiologists, chemists, enologists,
computational biologists, sociologists, economists, and the
grape industry
• 12 research institutions Dr. Bruce Reisch
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VitisGen Project
Genotyping
Genotyping-by-Sequencing (GBS)
Single Nucleotide Polymorphism (SNP)
Ultimate Goal: 50,000 SNPs/Population
43,971 SNPs have been identified.
A consensus map of 3,825 SNPs has
also been developed.
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Integration of Genetic Maps to construct
a high-resolution map with both SSR and
SNP markers using JoinMap 4.1 software
Genotyping: Genotyping-by-Sequencing (GBS)
Single Nucleotide Polymorphism (SNP)
Simple Sequence Repeat (SSR)
43,971 SNPs have been identified
A consensus map of 3,825 SNPs
A consensus map of 411 SSRs
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407 SSRs 1,665 SNPs
Total: 2,072 markers
Consensus Map
R/QTL software using a 4-way cross format
for composite interval mapping (CIM)
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There are only 4 gaps larger than 10 cM. JoinMap 4.1
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Adaxial Side Abaxial Side
Cabernet
Sauvignon
Cabernet
Sauvignon Norton Norton
Downy Mildew
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-20
0
20
40
60
80
100
120
0 50 100 150
Vis
ual
Microscope
r= 0.94
1
Norton Cabernet
Sauvignon
3ml Visual rating
Laboratory Assay
Field Assay
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Population Size 159
Repetitions 6
Duration 9 DAI
Downy Mildew
2014 Lab
2014 Field
2015 Lab
2015 Field
1 2 3 4 5
1 2 3 4 5
1 2 3 4 5
1 2 3 4 5
2014 lab 2015 lab 2014 Field 2015 Field
2014 lab 1.00 0.69 0.66 0.63
2015 lab 1.00 0.57 0.59
2014 Field 1.00 0.79
2015 Field 1.00
Strong correlations were observed
among data sets (Spearman
correlation coefficient = 0.57 to 0.79)
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Downy Mildew
Resistance (Rpv 25)
*LOD value of 16.4 explaining 33.8% of
the total phenotypic variation flanked
by markers VVCS1H077H166R1-1
(56.6 cM) and UDV737 (60.9 cM).
*Three SNP-trait association were
detected between the two flanking
SSR markers, further reducing the
interval distance to 0.7-2.3 cM.
Linkage Group 18
0
2
4
6
8
10
12
14
16
18
LO
D
2014 Lab
2015 Lab
2014 Field
2015 Field
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Norton Cabernet
Sauvignon
Botrytis Bunch Rot Botrytis cinerea
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Botrytis Bunch Rot Botrytis cinerea
Tropical Plant Pathology (2015) 40: 279
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Population Size 158
Repetitions 8
Duration 10 DAI
Botrytis Bunch Rot
0
10
20
30
40
50
60
70
80
90
100
1 6
11
16
21
26
31
36
41
46
51
56
61
66
71
76
81
86
91
96
101
106
111
116
121
126
131
136
141
146
151
156
Dis
ease
sev
eri
ty (
%)
Genotypes
Year 2016
0
10
20
30
40
50
60
70
80
1 5 9 13 17 21 25 29 33 37 41 45 49 53 57 61 65 69 73 77 81 85 89 93 97 101105109
Dis
ease
Sev
erit
y (%
)
Genotypes
Year 2015
1 2 3 4 5
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Botrytis Bunch Rot
Resistance
*LOD value of 7.1 explaining 18.4% of
the total phenotypic variation flanked
by markers VMC6F1 (42.7 cM) and
VMC3B10 (46.9 cM).
*Four SNP-trait association were
detected between the two flanking
SSR markers, , further reducing
the interval distance to 0.3-1.9 cM.
0
1
2
3
4
5
6
7
8
0 20 40 60 80 100
LO
D
2015 Lab 2016 Lab
Linkage Group 2
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Norton x Vignoles
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1. GBS data can be used to saturate the grape genome
with SNPs in a pseudo-testcross population.
2. SNPs and SSRs can have complementary roles: first, to
identify genome regions associated with traits of
interest using SNPs, and second, to perform marker-
assisted selection using SSRs.
3. The overall goal of this program is to provide molecular
genetic support to expedite a Norton grape breeding
effort with the ultimate goal of developing improved
cultivars well adapted to Missouri conditions.
Summary
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North American Grape Breeders Conference Arkansas, California, Florida, Georgia, Minnesota and New York
2015 Cornell University; 2017 UC Davis
2019 Missouri State University
August 15-16, 2019
Field Day – Thursday Missouri State Fruit Experiment Station, Mtn. Grove. MO
MSU breeding vineyard (VitisGen II) & winery; Virus cleaning network
St. James Winery, St. James, MO
Research Presentation – Friday Bond Learning Center, Springfield, MO
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VVCS1H077
H16R1-1
24.6 (Mb)
UDV305
24.9 (Mb)
UDV737/GF18-14
26.1 (Mb)
VMC7F2/GF18-08
26.9 (Mb)
Rpv3
1. Di Gaspero et al. (2012) studied the selective sweep in Rpv3 using its flanking
SSR loci UDV305 and UDV737. Seven different haplotypes were generated;
however, none of the haplotypes were present in ‘Norton’/’Cynthiana’.
2. The Rpv3 related SSR markers including UDV305, UDV108, UDV112 and VMC7F2
didn’t show polymorphism in Norton.
3. Downy mildew resistance in ‘Norton’ is most likely due to the presence of a new
locus within the shared region of Rpv3 (UVD305 and UDV737), or in the unique
region between markers of VVCS1H077H16R1-1 and UDV305.
Linkage Group 18
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Acknowledgements
Cornell University
Bruce Reisch
Lance Cadle-Davison
VitisGen I&II
UC Davis
Andy Walker
Li-Ling Chen Surya Sapkota
Rayanna Bailey Jacob
Schneider
Bryce Bentley Will Knuckles
Sadie Land
Funding Sources
USDA-NIFA-NLGCA Program
USDA-NIFA-AFRI Competitive Grant
Univ. Missouri
Jim English
Andy Thomas
Missouri Department of
Agriculture
E & J Gallo Winery, Modesto, CA
Missouri Grape & Wine Board