Transcript Need for a Public Doubled Haploid Facility

Need for a Public Doubled
Haploid Facility ?
Thomas Lübberstedt, Candy Gardner, Mike Blanco,
Uschi Frei, Elisabeth Bovenmyer
Structure
- DH Production in Maize: How ?
- Why (Doubled) Haploids ?
- Who uses DHs ?
- Need for a Public DH Facility ?
Technologies
1) Haploid production
- In vitro versus in vivo
- Maternal versus Paternal
2) Haploid identification
- Markers: Morphological or molecular
- Cytological / flowcytometry
3) Genome doubling
- Colchicine
- Other
In vitro: Anther Culture
In vivo Haploid Induction
F1
X
X
+
+
Paternal
Inducer (ig)
Maternal
Inducer
X
,
,
,
,
F2
,
,
Workflow & Bottlenecks Maternal DH Induction
Step
Challenge
Induction cross
- Inducer availability
- Induction rate
Haploid (embryo) selection - Visual scoring
- Background effects
- Inducer DNA introgression ?
Genome doubling
- Toxic Colchicine
- Alternative procedures
- Background effects
Selfing of double haploids
- Chimera -> partial fertility
Identification of Maternal Haploids
Donor
Inducer
Endosperm
Embryo
Outcrossed or
self-pollinated
Lethal
H embryo
F1 embryo
Geiger 2009
Comparison DH Approaches in Maize
Approach
Strength
Weakness
In vitro
- No need of inducer
- Low induction rate
- Genotype dependency
- Need of tissue culture
In vivo – paternal
- Simple inheritance
- cms conversion
- Low induction rate (2%)
In vivo – maternal
-Limited genotype
- Background effects
dependency
-Complex inheritance
- Induction rate (10%)
Why (Doubled) Haploids ?
Speed in Line Development
Founder line 1
Selfing
x
F1
Founder line 2
DH
Inbred line
Inbred line
Haploid Induction & Doubling of
Allelic Diversity Materials
Andrew Smelser et al., Plant Introduction Station, Ames
QTL MAPPING
Type of
Population
Strength
Weakness
F2:3
- Speed of production
- d and a estimates
- Heterogeneous families
RIL
- Homogeneous families
- Power of QTL detection
- Slow production
DH
- Speed of producing
homogeneous families
- Power of QTL detection
- Laborious production process
- Lower recombination (>RIL)
BC
- Speed of production
- Heterogeneous families
- a and d confounded
Trait / Gene Stacking
Line 1
Line 2
Probability for Fixation of Target Genes
X
No. of genes
F2
DH
1
0.25
0.5
2
0.0625
0.25
4
0.004
0.0625
8
0.00002
0.004
16
0.00000000002
0.00002
or
Selfing (F2)
DH Induction
Goal: Fixation of target alleles
Usefulness in Line Development
Seitz 2005
Rapid Line & Population Improvement
Gordillo & Geiger 2008
Conclusions
- Homozygous lines are obtained in short time
- Maximal genetic variation among lines: increased selection gain
- Reduces population size for gene pyramiding substantially
- Increases power for QTL or QTP detection
- Increases labor for line development – depending on technology
Who uses DHs ?
DH Application in Plant Breeding
Maize: All major breeding companies (in vivo DH induction)
Other species (Tuvesson et al.): Rye, Oat, Triticale, Potato, Cabbage
Tuvesson et al. 2007, EU-COST
Need for Public DH Facility ?
Pro and Contra Public DH Facility (Maize)
Contra:
- Inducers are available in maize
- Procedures are published
-> Everyone can do it
Pro:
- Users might need DH materials only once or once in a while
- Induction procedure requires know how (analogy to transformation)
- Technology advance might make procedure more accessible
- Possible platform for expansion into other species
=> Research opportunities in maize and other species
And: Training of future plant breeders !! Technology & Application of DHs
Technological Challenges
Questions relating to inducer:
- Alternative/improved marker systems
- Increased induction rate
- Adaptation to US climate / photoperiod
- Genetics / biology of induction mechanism
Colored Seed
Partial
Fertile
Questions relating to genome doubling:
- Genetic background effect ?
- Optimized procedure (with colchicine)
- Procedure without colchicine ?
- Increased doubling / success rate
Smelser et al. 2009
Conclusion
Successful example of research oriented service facility at ISU:
http://www.agron.iastate.edu/ptf/index.aspx
New DH facility at ISU will soon be launched, if interested contact:
Thomas Lübberstedt
515-294-5356
[email protected]
Collaborators:
Candy Gardner, Mike Blanco, Uschi Frei
Costs ?
Service
a) Haploid Seed Induction
a) Selected Haploid Seed
a) Doubled Haploid Seed Production
(5-20 kernels)
a) Doubled Haploid Line (200 kernels)
- Summer / Winter pricing
- GM seed ?
On Campus Fee Off Campus Fee