Factors Influencing the Composition/Metabolic Profile of Food … · 2020. 4. 5. · Plant Breeding...
Transcript of Factors Influencing the Composition/Metabolic Profile of Food … · 2020. 4. 5. · Plant Breeding...
Factors Influencing the Composition/Metabolic Profile of Food from Plants
Sherry Flint-Garcia, PhDUSDA Agricultural Research [email protected]
Conflict of Interest Statement
As a federal employee, I have no conflicts of interest to declare.
Outline
Domestication Plant Breeding and Biotechnology Maize Case Study
– Domestication and Diversity– Genetics of Kernel Composition– Composition and Metabolites
Appendix Slides– Overview of Genetics, Terms, and Methods
1. Domestication
Domestication: “Adaptation of plants and animals to intimate association with human beings”
Domestication Syndrome
Domesticates:– More robust plants– Synchronous flowering– Increased apical dominance– Non-shattering– Loss of seed dormancy– Larger fruits/grains
Peanut
Corn
Rice
CoffeeSoybean
Hops
Pistachio
Sorghum
DomesticatesProgenitors
USDA-ARS
Wheat
Complex evolutionary history due to polyploidizationand distinct market classes
USDA-ARS
Einkornwheat
Emmer wheat
Breadwheat
Durum (pasta) wheat
Ae. speltoides (BB)
T. tauschii(DD)
T. turgidum(AABB)
T. urartu (AA)
T. spelta(AABBDD)
T. aestivum(AABBDD)
Tomato
Details of domestication are unclear: likely Ecuador/Peru and Mexico. “Pre-domesticate” was a cherry tomato. Brought to Europe by Columbus or conquistadors <1544. Model system for genetic studies.
USDA-ARSPI 647555
USDA-ARS
PI 347239PI 634844
Wild Ancestor: Solanumpimpinellifolium Pre-domesticate:
S. lycopersicumssp cerasiforme
Domesticate: S. lycopersicumssp lycopersicum
2. Plant Breeding and Biotechnology
Plant Breeding: “The art and science of plant improvement”Biotechnology: “The use of living systems and organisms to develop or
make useful products”
First Plant Breeders
Thousands of years ago– Likely women who selected plants that they liked.
Hundreds of years ago– Farmers had their favorite “family” variety = “heirloom.”
1926 Pioneer Hi-Bred was founded– First commercial hybrid corn seed company.
Plant Breeding
Selection (artificial) of plants with desirable traits.– Often done without knowledge of the genes controlling the trait(s).
Decreasing Unfavorable Alleles.– Susceptibility to diseases and insects, susceptibility to drought,
unadaptedness. Increasing Favorable Alleles.
– Increased yield and better quality, resistance to disease, insects and environmental stresses.
This process has typically produced safe food for over 100 years.
Plant Breeding
• Changing the phenotypic mean of a population; two methods
Select the “best”
individuals to form
the next generation
Original population
200
Progeny population
218
Parent 1Low yield
Insect resistant
F1
Parent 2High yieldSusceptible
F2
Select the “best” individuals to form the next generation
New Inbred LineHigh yieldInsect resistant
Plant Biotechnology
• Requires knowledge of the gene underlying the trait.• Gene(s) excised from donor species, combined with promoter of
choice to drive gene expression.• The construct is transformed into recipient plant.• Backcrossed six times to recover 99.2% recurrent parent.
Commercial Variety
Donor Plant
NewVariety
Traditional Plant BreedingDNA is a strand of genes, much like a strand of pearls.Traditional plant breeding transfers many genes at once.
Desired gene Many genes are transferred
Plant BiotechnologyUsing plant biotechnology, you can add a single gene to the strand without the extra baggage (linkage drag).
Donor Commercial Variety
Desired gene Desired gene
× +A single gene is transferred
Improved Commercial
Variety
3. Maize Case Study
What Can We Learn From Maize Diversity Using Breeding, Genetics, and Genomics?
Maize is an important crop! Domestication and Diversity Inbred Lines Teosinte, the Wild Ancestor
Maize Diversity Project
www.panzea.org
Maize diversity is greater than the difference between human and chimps
Silent Diversity; Tenallion, et al. (2001) PNAS
1.42% 0.09%1.34%
Maize Domestication
Domesticated from Zea mays ssp. parviglumis ~9,000 years ago in SW Mexico.Intermediate form of landraces, i.e., populations adapted to specific microclimates
and/or human uses.
Artificial Selection
Teosinte (ssp parviglumis) Inbred LinesLandraces
The Maize Genome
Ten chromosomes. Ancient tetraploid – whole genome duplication ~ 5-12 mya. B73 is the reference inbred for the genome. 32,500 predicted genes in B73 – more likely ~40,000. Nearly 85% of the B73 genome sequence is annotated as
transposable elements. B73 may capture only ~70% of the low-copy gene fraction of all maize
inbred lines.
B73
Schnable, et al. (2009) Science
Genome-Wide Diversity Maize HapMap.v1, HapMap.v2, HapMap.v3
– 83 million SNPs Two historically important inbred lines in breeding (B73 and Mo17)
differ by more than 20 million SNPs! Related species exhibit synteny (same gene order) along
chromosomes. However, we see lots of non-collinearity in maize!
Gore, et al. (2009) Science; Chia, et al. (2011) Nature Genetics; Bukowski, et al. (2018) GigaScience; Fu and Dooner (2002) PNAS
GCCATC ---CGA
GCGATCGGGCGA
SNP In/Del
Domestication Genes in Maize
• teosinte branched 1 – plant branching• teosinte glume architecture 1 - hard fruitcase
• Whole genome analysis of HapMap.v2– Stronger selection during domestication than improvement– > 1000 genes experienced selection
Wang, et al. (2005) Nature; Hubbard, et al. (2002) GeneticsHufford, et al (2012) Nature Genetics
Impact of Artificial Selection on Diversity
Teosintes
MaizeLandraces
MaizeInbred Lines
Unselected (Neutral) Gene Domestication Gene Improvement Gene
Artificial Selection
Plant Breeding
Domestication
98% (~49,000)maize genes
2% (~1,000) maize genes Commercial Corn Breeding
?
Diverse Maize Inbred LinesNested Association Mapping (NAM)
Linking Phenotype to Genotype
Can we identify the genes controlling phenotypic variation?
QTL (Quantitative Trait Locus) Mapping– Identify genomic regions that contribute to variation– Very loose definition of a large region
Association Mapping– Test specific genes for their involvement in the trait– Much more precise way to test (if done correctly!)
Association Panel
302 Inbred Linesfrom around theworld
Flint-Garcia, et al. (2005) Plant J.
0.1
NC33
B115
I137TN
81-1
MEF 156-55-2
IL677A
Ia5125
IA2132
IL101
F2EP1
F7
CO255
NC366
B52
SC213R
NC238
GT112
Mp339
GA209
D940Y
T232
U267Y
CI28A
B2
F2834T
Mo24WMS1334
IDS28
I-29
SA24
4722
SG18Sg1533
IDS91IDS69
F6
F44
Ab28A
CML328
Oh603
A441-5
CML323
SC55
NC264NC370
NC320
NC318
NC334
NC332
CML92
CML220
CML218A272
Tzi9
CML77
TZI10
CML311
CML349
CML158Q
CML154Q
CML281
CML91
parvi-14
parvi-36
parvi-03
parvi-49
parvi-30
NC356
TX601
NC340
NC304
NC338
NC302
NC354TZI18
A6
Ki44
Ki43
Ki2007Ki21
Ki2021
Ki14
CML238CML321
CML157Q
CML322
CML331CML261
CML341CML45CML11
CML10
Q6199
CML314
CML258
CML5CML254
CML61
CML264
CML9
CML108
CML287
Tzi11
CML38
CML14
SC357L578
T234
N6E2558W
K64
CI64
CI44
CI31ANC230
CI66
K55
R109BMoG
L317
NC232
VaW6
CH701-30
Va85
CI90C M14
H99
Va99 PA91
Ky226
H95
Oh40B
VA26
Pa762
A619
Va22
Va17
Va14 Va59
Va102
Va35
T8
A654
Pa880
SD44
CH9
Pa875
H49
W64A
WF9
Mo1W
Os420
R168
38-11
NC260
Mo44
CI21E
Hy
A661
ND246
WD
A554
CO125
CO109
B75
DE-3
DE-2
DE1
B57
C123
C103
NC360
NC364
NC362
NC342
NC290A
NC262
NC344
NC258
CI91B
CI187-2
A682
K4
NC222
NC236
CI3A
K148
Mt42
MS153
W401
A556
B77
CM37
CMV3
W117HT
CM7
CO106
B103
R4
Ky228
B164
Mo46
Mo47
Mo45
Hi27
Yu796-NS
I205
Tzi16Tzi25
B79
B105
N7A
SD40
N28HT
A641
DE811
H105W
A214N
H100
A635
A632
A634B14A
H91B68
B64
CM174
CM105B104
B84
NC250
H84
B76
B37
N192
A679
B109
NC294
NC368NC326
NC314
NC328 R229
A680
NC310
NC324
NC322NC330
NC372
B73Htrhm B73NC308
NC312 NC306
NC268
B46B10
A239
C49AC49
A188
W153R
A659
R177
W22
W182B
CI-7
33-16
4226
NC348
NC298
NC352
NC336
NC296
NC346NC296A
NC292
SWEET
POPCORNTROPICAL-SUBTROPICAL
STIFF STALK
NON STIFF STALK
MIXED Based on 89 SSR loci
MO17
Oh43E
teosinte
NC300
NAM Founders
25 Diverse Lineschosen to maximize diversity
0.1
NC33
B115
I137TN
81-1
MEF 156-55-2
IL677A
Ia5125
IA2132
IL101
F2EP1
F7
CO255
NC366
B52
SC213R
NC238
GT112
Mp339
GA209
D940Y
T232
U267Y
CI28A
B2
F2834T
Mo24WMS1334
IDS28
I-29
SA24
4722
SG18Sg1533
IDS91IDS69
F6
F44
Ab28A
CML328
Oh603
A441-5
CML323
SC55
NC264NC370
NC320
NC318
NC334
NC332
CML92
CML220
CML218A272
Tzi9
CML77
TZI10
CML311
CML349
CML158Q
CML154Q
CML281
CML91
parvi-14
parvi-36
parvi-03
parvi-49
parvi-30
NC356
TX601
NC340
NC304
NC338
NC302
NC354TZI18
A6
Ki44
Ki43
Ki2007Ki21
Ki2021
Ki14
CML238CML321
CML157Q
CML322
CML331CML261
CML341CML45CML11
CML10
Q6199
CML314
CML258
CML5CML254
CML61
CML264
CML9
CML108
CML287
Tzi11
CML38
CML14
SC357L578
T234
N6E2558W
K64
CI64
CI44
CI31ANC230
CI66
K55
R109BMoG
L317
NC232
VaW6
CH701-30
Va85
CI90C M14
H99
Va99 PA91
Ky226
H95
Oh40B
VA26
Pa762
A619
Va22
Va17
Va14 Va59
Va102
Va35
T8
A654
Pa880
SD44
CH9
Pa875
H49
W64A
WF9
Mo1W
Os420
R168
38-11
NC260
Mo44
CI21E
Hy
A661
ND246
WD
A554
CO125
CO109
B75
DE-3
DE-2
DE1
B57
C123
C103
NC360
NC364
NC362
NC342
NC290A
NC262
NC344
NC258
CI91B
CI187-2
A682
K4
NC222
NC236
CI3A
K148
Mt42
MS153
W401
A556
B77
CM37
CMV3
W117HT
CM7
CO106
B103
R4
Ky228
B164
Mo46
Mo47
Mo45
Hi27
Yu796-NS
I205
Tzi16Tzi25
B79
B105
N7A
SD40
N28HT
A641
DE811
H105W
A214N
H100
A635
A632
A634B14A
H91B68
B64
CM174
CM105B104
B84
NC250
H84
B76
B37
N192
A679
B109
NC294
NC368NC326
NC314
NC328 R229
A680
NC310
NC324
NC322NC330
NC372
B73Htrhm B73NC308
NC312 NC306
NC268
B46B10
A239
C49AC49
A188
W153R
A659
R177
W22
W182B
CI-7
33-16
4226
NC348
NC298
NC352
NC336
NC296
NC346NC296A
NC292
SWEET
POPCORNTROPICAL-SUBTROPICAL
STIFF STALK
NON STIFF STALK
MIXED Based on 89 SSR loci
MO17
Oh43E
teosinte
NC300
NC350
P39
M37WHP301CML277
B97
CML103CML69
CML52
CML228
CML247
CML332
IL14H
Ky21
Ki11
Ki3
MS71
Mo18W
OH7B
M162W
Tx303
TZI8CML333
NC358
OH43
Flint-Garcia, et al. (2005) Plant J.
Nested Association Mapping
NAM combines the statistical power and mapping resolution from twodifferent QTL mapping methods– Explore diversity from around the
world Used to study many different
traits in maize
25 DL SSD
. . .
B97
CML103
CML228
CML247
CML277
CML322
CML333
CML52
CML69
Hp301
Il14H
Ki11
Ki3
Ky21
M162W
M37W
Mo18W
MS71
NC350
NC358
Oh43
Oh7B
P39
Tx303
Tzi8
⊗ ⊗ ⊗
⊗ ⊗ ⊗
⊗ ⊗ ⊗
Asso
ciat
ion
Linkage = QTL mapping
Yu, et al. (2008) Genetics McMullen, et al. (2009) Science
NAM Kernel Composition
Jason CookStarch
Amylose
Amylopectin
Fiber
Oil
Fatty Acid Profiles
Protein
Zeins
Amino Acid Profiles
Cook, et al. (2012) Plant Phys.
Kernel Composition QTL Mapping in Maize
0
10
20
30
40
50
60
70
80
LOD
StarchProteinOil
Chr. 1 2 3 4 5 7 8 9 10
“QTL for kernel oil content somewhere in here.”
Cook, et al. (2012) Plant Phys.
Genome Wide Association (GWAS)
1.6 Million HapMap.v1 SNPs projected onto NAM
0.0
10.0
20.0
30.0
40.0
50.0
60.0
70.0
80.0
90.0
100.0
- 500,000,000 1,000,000,000 1,500,000,000 2,000,000,000
Physical Distance (bp)
GWAS - Oil
RM
IP
Getting closer to saying “This genecontrols oil content”Cook, et al. (2012) Plant Phys.
Teosinte - The Wild Ancestor
Development of Teo Introgression Libraries
B73 parviglumis
BC1 BC2 BC3 BC4
F1 Hybrid: B73×parviglumis
Backcrossing of the F1 with B73 to create BC1, BC2, BC3, BC4;Each backcross removes 50% of the teosinte genome.
Liu, et al. (2016) Plant Genome
Kernel Composition QTL in Teosinte
Karn et al. (2017) G3
Avi KarnTeosinte NILs
Chr.1 2 3 4 5 6 7 8 9 10
Allelic effects (% starch)
Cook et al. (2012) Plant Phys. & Karn et al. (2017) G3
Teosinte NILs
NAM RILs
Chr.1 2 3 4 5 6 7 8 9 10
Application: Reintroduce Variation
Biological hypothesis: A loss of genetic variation results in a loss of phenotypic variation.
Breeding hypothesis: We can improve modern corn by introducing variation.
Sucrose
UDP Glucose
ADP Glucose
Amylose Amylopectin
Sh1 : sucrose synthase
Sh2, Bt2 : ADP-glucose pyrophosphorylase
Su1 : DBEAe1 : SBE-IIB
Wx1 : GBSS
Selected Genes in Maize Starch Pathway; Whitt et al. (2002)
Gene ID Gene Name Possible Target TraitGRMZM2G348551 Su2; Sugary 2 StarchGRMZM2G394450 Ivr1; Invertase 1 StarchGRMZM2G089836 Ivr2; Invertase 2 StarchGRMZM2G110175 Bm1; Brown midrib 1 StarchAC196475.3_FG004 Bm3; Brown midrib 3 StarchGRMZM2G098298 Ccp1; Cysteine protease 1 ProteinGRMZM2G138727 Zp27; 27-kDa zein protein Protein & Amino AcidsGRMZM2G087612 SDP1; Sugar dependent1 Oil
Hufford et al. (2012) Nature GeneticsSchnable and Freeling (2011) Plos One
Do Teosinte Alleles Have Value for Improving Today’s Corn?
Traits that were relevant 9000, 1000, or 100 years ago may not be as relevant today.
Therefore the alleles selected 9000, 1000, or 100 years ago that persist in modern germplasm may not be optimal today.
Conclusions: Impact of Selection in Maize
Selection on the genome – Thousands of years during domestication– Over one hundred years during plant breeding– 25+ years during plant biotechnology
Modification of seed composition– Traditional breeding– Biotechnology
Never identified a deadly/toxic variety of corn– Despite tremendous genetic and phenotypic variation– Long history of safe use
Surveys of Metabolites in Diverse Maize Germplasm
27 Nested Association Mapping (NAM) founder inbreds27 Landrace (LR) inbreds created by selfing landracesBoth sets crossed with B73 to make hybrid seed
B73 Genome Inbred
Genetic Relationships Among the Maize Lines
Text in this font and color• First level bullet
− Second level dash
NAM Hybrids
Landrace Hybrids
LandracesNAM Inbreds
Venkatesh et al. (2015) J. Agric. Food Chem.
Compositional Variation in Diverse Maize
Venkatesh et al. (2015) J. Agric. Food Chem.
Compositional Variation in Diverse Inbred Lines
Wide range inproximatesamong inbredlines
Total Fat Total Carbohydrate
Venkatesh et al. (2015) J. Agric. Food Chem.
Compositional Variation in Diverse Inbred Lines
Two folddifference in some fatty acidsamong inbred lines
C18:3 (Linolenic Acid) C18:1 (Oleic Acid)
Venkatesh et al. (2015) J. Agric. Food Chem.
Inbred lines versus landraces: Gamma-Tocopherol
Nearly six-folddifferencein gamma-tocopherolacross inbred lines andlandraces
LandracesInbred Lines
Venkatesh et al. (2015) J. Agric. Food Chem.
Inbred lines versus landraces: Beta-Carotene
Huge rangeof variationfor beta-carotene
LandracesInbred Lines
Venkatesh et al. (2015) J. Agric. Food Chem.
Metabolite Profiles by GC-MS and NMR
Same samples as previous study GC-MS and NMR were used to examine metabolite profiles 675 total metabolites collected by GC-MS
– 504 met QC checks; 174 were annotated 33 total metabolites collected by NMR
– 17 amino acids, 7 organic acids, 5 sugars, 4 others
Partial least-squares discriminant analysis (PLS-DA) of the GC-MS data
Venkatesh et al. (2016) J. Agric. Food Chem.
Another source of variation: Xenia Effect (Pollen)
Corn is a cross-pollinated species (pollen travels in the air)
R-locus
Y-locus
Sh2-locus (sweet corn)
Ornamental Corn
Xenia Effect (Pollen Donor) on Amino Acids
β γα family
δ
Flint-Garcia et al. (2009) Theor App Genet Unpublished data, Flint-Garcia (USDA) and Angelovici (University of Missouri) labs
EmbryoEndosperm
Zeins
Zein Profiles Xenia ExperimentZeins are the storage proteins in the endosperm.
To Summarize
Maize is extremely genetically diverse. Maize is extremely phenotypically diverse.
– Plant-level metabolite level Genetics, Environment, GxE contribute to phenotypes. Many genes contribute to grain composition. Even pollen source affects grain composition.
To Summarize
For geneticists and breeders, the more variation the better!
When a farmer plants their corn, every hybrid seed is genetically the same. When the farmer harvests the seed, every seed is different.
References Bukowski, et al. (2018) GigaScience; 7:1–12 Chia, et al. (2011) Nature Genetics 40:803-
807 Cook, et al. (2012) Plant Phys. 158:824-834 Flint-Garcia, et al. (2005) Plant J. 44:1054–
1064 Flint-Garcia, et al. (2009) Theor App Genet
119:1129-1142 Fu and Dooner (2002) PNAS 99:9573–9578 Gore, et al. (2009) Science 326:1115–1117 Hubbard, et al. (2002) Genetics 162: 1927–
1935 Hufford, et al (2012) Nature Genetics 44:808-
811
Karn, et al. (2017) G3 7:1157-1164 Liu, et al. (2016) Plant Genome 9 Schnable, et al. (2009) Science 326:
1112−1115. Schnable and Freeling (2011) PLOS ONE 6:
e17855. Tenaillon, et al. (2001) PNAS 98:9161-9166 Venkatesh, et al. (2015) J. Agric. Food
Chem. 63:5282−5295 Venkatesh, et al. (2016) J. Agric. Food
Chem. 64:2162−2172 Wang, et al. (2005) Nature 436:714–719 Whitt, et al. (2002) PNAS 20:12959–12962
Appendix Slides: Overview of Genetics & Breeding
Genetics: “The study of heredity and the variation of inherited characteristics”
Breeding: “The art and science of the genetic improvement of plants”
Genetics Terms
Genotype - The genetic constitution of an individual organism Phenotype - The set of observable characteristics of an individual
resulting from the interaction of its genotype with the environment Trait - A genetically determined characteristic Allele - An alternative form of a gene Genetic Variation - Naturally occurring genetic differences among
individuals of the same species
Genetics 101
TranscriptionDNA RNA
TranslationProtein
Glutamine|
Glycine|
Leucine|
Threonine |
Histidine |
Arginine|
Serine
Nucleotides
CAGGGACUGACUCAUCGAUCA
Sugar-phosphatebackbone
(a) primary structure -amino acids in
linear order
Base Pairs
(b) secondary structure – helices or sheets of amino acids(c) tertiary structure – folding of the protein into a shape(d) quaternary structure – multiple proteins form a complex
Where Does Genotypic Variation Come From?
Mutations– Point mutations – Change or delete base pairs– Chromosomal – Polyploidy and larger scale mutations
Transposable Elements – “Jumping Genes”– Can completely disrupt gene function
Hybridization– Two different genomes come together
Recombination – Shuffling of genes during meiosis
AATCGCGTTAAATCCCGTTA
DNA Sequence Variation
SNP: Single Nucleotide Polymorphism
InDel: Insertion/Deletion
Where Does Phenotypic Variation Come From?
Genotype– Why fraternal twins look different
Environment– Why identical twins look different
Genotype x Environment– Complex interactions between
genetics and the environment
How Does Variation Change?
Usually variation (allele frequencies) is balanced, i.e., “in Hardy-Weinberg Equilibrium.”
Genetic Drift– Sampling error – Rarer alleles can disappear completely and thereby reduce
genetic variation. Selection – Natural or Artificial
– Increased survival and reproduction of some individuals Gene Flow
– Migration of individuals, bringing new alleles together
Types of Variation
Qualitative = Discrete Quantitative = Continuous
dwarf 8
Male sterile 8
Albino seedling
Cob Color
NativeRootworm Resistance
Seed Composition
Carotenoid (proviatmin A)
Yield
Quantitative Trait Locus (QTL) Mapping
• “Genome Scan”• Identify genomic regions that contribute to variation and estimate QTL
effects Parent 1
F1
Parent 2
F2
u162
2
u155
2
b227
7
m23
1
b224
8
b122
5
b207
7
b152
0
0
1
2
3
4
56
7
8
9
0 10 20 30 40 50 60 70 80 90 100
110
120
130
140
Chr 1, Position (cM)
LOD
Sco
reGenotypePhenotype
Composite Interval Mapping
Association Analysis
“Genome Scan” Utilizes natural populations
− Exploit extensive ancestral recombination
1.3m
1.5m
1.4m
1.8m
2.0m
2.5m
T
C
C
T
T
C
A
G
G
A
G
G
G
G
T
T
T
G
A
A
A
C
A
A
A
A
A
G
G
G
Line 1
Line 2
Line 3
Line 4
Line 300…
Gene ABC
Association Analysis
“Genome Scan” Utilizes natural populations
− Exploit extensive ancestral recombination
1.3m
1.5m
1.4m
1.8m
2.0m
2.5m
T
C
C
T
T
C
A
G
G
A
G
G
G
G
T
T
T
G
A
A
A
C
A
A
A
A
A
G
G
G
Line 1
Line 2
Line 3
Line 4
Line 300…
Gene ABC
Nested Association Mapping (NAM)
Genome scanStructured populationHigh powerLow resolutionAnalysis of 2 alleles
Association Mapping Candidate gene testing
Unstructured populationLow power
High resolutionAnalysis of many alleles
Nested Association Mapping (NAM)Structured families nested within an unstructured population
High powerHigh resolution
Analysis of many alleles
Linkage (QTL) Mapping