Development and Characterization of Maize-Teosinte Introgression Libraries
Sherry Flint-GarciaUSDA-ARS Columbia, MO
OutlineIntroductionDevelopment of Teo NIL LibrariesApplications of Teo NIL LibrariesLofty Ideas
Evolutionary Genetics of Maize
Domesticated from Zea mays ssp. parviglumis, existed/exists in an intermediate form of landraces
Single domestication event in Mexican highlands
6,000~9,000 years ago
Selection
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
GENOMICS Nested Association Mapping (NAM)
BREEDINGGermplasm Enhancement
of Maize (GEM)
Teosinte Introgression LibrariesGENETICS
Impact of Artificial Selection
Development of Teo NIL Libraries
B73 × teosinte(parviglumis)
10 accessions
F1 :B73 × teosinte
teosinte BC1 BC2 BC3 BC4B73 F1
Library Development161 BC4S2 NILs: 2 libraries (done in 2009)643 BC4S2 NILs: 8 libraries (done in 2008) 83 BC4DH NILs: duplicated library887 Near Isogenic Lines (NILs)
Z031E0035
Z035E0012
Illumina GoldenGate
768 SNP assay
Introgression Stats# Avg. # Avg. % Avg. % Avg. %
Population Lines Regions Teosinte Het. TeozygousCoverage
Ames 21785 96 2.5 4.5 2.9 1.6 4.3 Ames 21786 87 2.3 4.4 2.9 1.5 3.8 Ames 21789 93 2.5 3.9 2.9 1.1 3.6 Ames 21814 77 2.1 3.8 2.5 1.3 2.9 Ames 21889 91 2.7 4.7 3.3 1.3 4.2 Ames 21809 79 2.7 4.9 3.1 1.7 3.8 Ames 21812 82 2.1 4.6 3.1 1.5 3.8 PI 384065 53 2.5 4.2 2.4 1.7 2.2 PI 384066 64 2.2 4.0 2.5 1.5 2.6 PI 384071 82 2.3 4.1 2.6 1.5 3.3 PI 384071 - DH 83 1.4 1.9 0.1 1.8 1.6
Expected Genotype Ratios for BC4S2 (sibbed): 95% BB, 3% BT het., 1.5% TT
Library Coverage
c1 c2 c3 c4 c5 c6 c7 c8 c9 c10
10 maize-teosinte libraries804 BC4S2 NILs and 83 BC4DH NILsEach line:
2.3 chromosomal segments4.1% of the teosinte genome
3.3X genome coverage
KEY
0123456789
BC4S2 vs BC4DH Coverage
KEY
0123456789
c1 c2 c3 c4 c5 c6 c7 c8 c9 c10S2 DH S2 DH S2 DH S2 DH S2 DH S2 DH S2 DH S2 DH S2 DH S2 DH
. . 2 2 . . 3 1 7 2 . . 2 1 2 0 3 5 2 03 0 2 3 6 2 . . 6 2 . . 1 1 4 0 5 7 . .3 0 . . 7 3 3 2 7 2 . . . . 3 0 4 5 . .3 0 . . 6 2 3 0 . . . . 1 1 . . 4 5 2 03 0 3 3 6 2 . . . . 3 2 2 1 2 1 4 5 . .. . . . 6 2 2 0 5 0 4 2 2 1 1 1 5 5 . .. . 3 4 4 3 . . 5 0 . . 2 1 . . 4 3 3 03 0 3 4 4 3 . . . . 4 2 . . 1 2 . . . .. . 3 3 4 3 3 0 2 0 4 2 1 1 . . 3 3 . .. . 3 3 . . 2 0 3 0 3 2 2 1 1 2 4 3 1 0. . 2 3 5 4 3 0 . . . . 1 1 . . 3 1 . .1 0 2 3 5 4 . . 2 0 3 2 2 1 . . . . 1 02 0 2 2 . . 2 1 1 0 3 2 3 2 1 0 . . . .2 0 . . 6 1 4 1 1 0 4 2 4 2 1 0 . . 1 02 0 . . 7 1 3 1 1 0 . . 3 2 1 0 1 0 0 02 0 . . 7 1 4 1 1 0 4 2 4 2 1 0 2 1 1 02 0 3 2 . . 4 1 1 0 4 3 3 2 0 0 1 1 . .1 0 3 2 . . . . 2 0 3 2 2 2 2 0 . . 1 01 0 3 2 7 1 4 1 2 0 3 2 2 2 1 0 1 1 1 01 0 . . 8 1 . . 2 0 5 0 2 2 1 0 . . . .. . 5 1 8 1 4 1 3 0 4 1 . . 1 0 . . 2 02 0 5 2 . . 4 1 3 0 5 0 2 2 1 1 1 0 1 12 0 3 2 . . 4 1 3 0 . . 2 2 1 1 1 0 1 02 0 5 2 8 1 4 1 3 0 3 0 3 1 0 1 1 0 1 11 0 . . 8 1 4 1 2 1 . . 4 2 0 2 . . . .. . 5 2 . . 4 1 3 1 4 0 4 2 . . 1 0 . .2 0 5 1 8 1 3 1 3 1 4 0 3 2 0 1 2 0 2 1. . 5 2 7 1 . . 2 1 5 0 . . 1 2 2 0 2 12 0 . . 7 1 4 1 3 1 . . 3 1 0 2 1 0 2 1. . 5 2 . . 5 1 2 2 5 0 2 3 0 2 1 0 2 12 0 4 1 6 1 . . 3 2 5 0 4 3 . . 2 0 . .. . 2 1 7 2 5 1 3 2 7 0 3 2 . . 2 0 2 12 0 1 1 5 2 4 1 3 2 7 0 . . . . 1 0 2 13 0 3 2 . . . . . . . . 4 2 0 2 1 1 . .2 0 . . . . 5 1 4 2 5 1 4 2 1 3 . . . .3 0 . . 6 3 4 1 4 2 6 0 . . 1 3 . . . .2 0 3 2 . . 3 0 4 2 5 2 4 2 1 3 1 1 4 04 0 2 3 6 3 . . 4 2 6 2 3 1 1 3 1 1 4 04 0 3 3 5 4 5 1 4 2 6 2 4 1 1 4 1 1 . .4 0 2 2 . . 6 2 3 1 5 4 4 1 . . . . 4 03 0 3 2 6 3 . . 4 2 4 3 2 2 1 4 1 1 4 03 0 3 2 6 4 . . 3 0 6 2 2 2 1 4 1 0 3 03 0 3 2 6 4 . . . . 4 2 1 1 . . 1 1 . .. . 2 2 2 5 1 1 4 1 . . 0 1 4 4 2 0 2 13 0 3 2 6 4 1 1 4 1 5 2 . . 4 4 2 1 . .3 0 2 2 7 4 1 2 4 1 5 2 1 1 5 4 2 1 2 13 0 2 1 . . 1 1 4 1 5 2 5 3 2 2 4 1. . 2 1 6 4 . . 4 1 5 1 . . 2 2 . .3 0 3 1 6 4 1 3 4 1 5 1 5 4 2 2 4 13 0 3 1 6 6 3 3 4 1 4 0 5 4 . . 4 13 0 3 1 6 7 3 2 4 1 . . 5 4 3 23 0 3 1 6 7 3 3 4 1 2 0 4 4 2 22 0 3 0 5 6 3 3 3 1 . . 5 4 0 03 0 3 1 5 7 3 3 4 1 . . . . . .3 1 4 2 5 7 3 3 4 0 3 1 5 4 1 03 1 4 2 4 3 3 3 2 2 3 1 5 4 2 13 1 . . 4 3 . . . . 6 33 1 4 2 . . . . 2 1 4 23 1 3 1 4 1 3 2 2 2 4 33 1 4 2 . . 3 2 3 2 4 13 1 . . 4 2 3 3 . . 5 13 1 5 2 3 1 4 3 . . 6 2. . 5 2 2 1 3 3 2 2 6 22 3 4 2 2 1 . . 3 2 6 22 3 5 0 1 1 3 3 3 2 . .. . 6 3 1 2 1 2 3 1 . .. . 5 3 1 2 1 2 3 1. . . . . . 1 2 3 12 3 . . . . 1 3 2 12 3 3 3 . . 1 2 2 12 3 . . 3 5 . . 3 12 2 3 6 3 4 . . 6 1. . 2 6 2 4 3 3 . .. . 3 7 3 4 3 3 6 13 1 4 6 3 4 5 13 1 . . 3 4 5 0. . . . 3 4 5 1. . 4 7 4 4 6 13 2 3 7 4 4 5 13 2 5 6 . . 2 24 2 5 5 5 4 5 10 0 . . . . 6 04 2 4 4 5 4 . .5 1 8 3 3 2 6 06 1 8 3 3 2 7 05 1 6 3 7 06 3 . .6 3 . .5 3 7 04 3 7 04 2 7 0. . . .3 2 8 0. . . .3 2 7 1. . . .2 0 . .. . 6 23 1 5 2. . . .2 1 2 03 13 14 13 13 1. .5 14 14 15 1. .5 15 1. .5 12 1
Applications of Teosinte NILs
1. Empirical Genetics Questions1000 Selected Genes
What do these selected genes do?What traits were targeted by artificial selection
during domestication/breeding?Are selected genes important?
Auxin response factor, ARF1
0
0.01
0.02
1000 2000 30001 (bp)
Div
ersi
ty (π
)
InbredsTeosinte
Tillering/branching?
?Auxin-mediatedProtein
Degradation
Could be any trait!
2. Evaluate Allele Series
Examine the range of allele effects of maize versus teosinte.
Validate QTL identified by NAM.
Trait ValueB73
MaizeAlleles
TeosinteAlleles
A Brief DigressionNested Association Mapping (NAM)
Flowering Time
Ed Buckler, Jim Holland, Mike McMullen, et al
Buckler, et al. (2009) Science
Flowering Genetic ArchitectureAt least 39 QTL explain flowering architectureGenetic architecture of maize is very complex
and very different than Arabidopsis.
Buckler, et al. (2009) Science
Many QTL with small Effects
333 alleles significant at P = 0.05.
Only 7 alleles had more than 24 hour effect.
Buckler, et al. (2009) Science
-150
-100
-50
0
50
100
150
Freq
uenc
y of
QTL
12h
Significant QTL
24h 36h
Increase Flowering Time
Decrease Flowering Time
Additive Allelic Effects
Allele Series
69% of QTLs had both alleles with both positive and negative effects.
Buckler, et al. (2009) Science
1 39……………….………………. QTL……………………….……….
Additive Allelic Effects (days)
“Gene Stacking”
Large differences in parental lines (32 day span) are the product of stacking large numbers of modest effect QTL.
Buckler, et al. (2009) ScienceEarliest Latest
Back To Teosinte
Flowering Evaluation~850 NILs from 10 librariesDays to Anthesis (DTA) from:
Ithaca, NY (2 reps), Rebecca Nelson & Oliver OttIthaca, NY (2 reps), Buckler labRaleigh, NC (8 reps), Peter Balint-Kurti Madison, WI (1 rep), Loren TrimbleSt. Paul, MN (1 rep), Nathan SpringerColumbia, MO (4 reps)
TOTAL 18 reps
0102030405060708090
0 100 200 300 400 500 600 700 800 900 1000 1100 1200 1300 1400
0
10
20
30
40
50
0 100 200 300 400 500 600 700 800 900 1000 1100 1200 1300 1400
Teosinte Introgressions vs. NAM
Additive Effect(days)
Teosinte NILs
1.1
NAMPhoto-period Gene?
-1.01.6
-1.13.2
-2.64.2
-0.92.3 1.9
-2.91.3
-0.72.2
-2.32.4
-1.20.8
1.03.8
-4.12.2
-1.02.8
0.74.5
0.7 0.7
Sucrose
UDP Glucose
ADP Glucose
Amylose Amylopectin
sh1 : sucrose synthase
sh2, bt2 : ADP-glucose pyrophosphorylase
su1 : debranching enzymeae1 : SBE-IIB
wx1 : g.b starch synthase
Selected Genes in Maize Starch Pathway; Whitt et al. 2002
3. Reintroduce Genetic Variation
Biological hypothesis: A loss of genetic variation results in a loss of phenotypic variation.
Breeding hypothesis: We can improve modern traits.
Kernel Traits
0
10
20
30
40
50
60
70
80
Carbohydrate Protein Fat
Perc
ent o
f Ker
nel W
eigh
t
Teosinte (N = 11)Landraces (N = 17)Inbred Lines (N = 27)
Flint-Garcia et al. (2009) TAG
seed size
β γ
α family
δ
Zein Profile
Teosinte NIL Evaluations
Hibbard – Corn Rootworm
Resistance
Buckler – Flowering
Brutnell – ShadeAvoidance Response
Balint-Kurti – SCLB and GLS
Smith – Smut
Tracy – Germination
AgReliant – Agronomic
TraitsNelson – NCLB
Hoekenga – Iron Bioavailability
Harmon – Circadian
Clock
Dallo – Mal de Rio Cuarto Disease
Flint-Garcia –Starch, Protein, Oil
Zein ProfileAmino Acid Profile
Lofty Ideas
2010
AgronomicsFertilizerDensity
Mechanization
?
Select only on yield
ideotype
7000 BC
Teosinte Synthetic75% B73 (SS), 25% teosinte
Acknowledgements
www.panzea.org
NSF Maize Diversity Project
SyngentaAgReliant