need for a public doubled haploid facility ? thomas lübberstedt, candy gardner, mike blanco, uschi...
TRANSCRIPT
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 ?
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
Technologies
In vitro: Anther Culture
In vivo Haploid Induction
F1
X
, ,F2
, ,
X
Maternal Inducer
+
, ,
+X
Paternal Inducer (ig)
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
Endosperm
Embryo
H embryo F1 embryoLethalOutcrossed or self-pollinated
Donor Inducer
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 dependency- Induction rate (10%)
- Background effects-Complex inheritance
Why (Doubled) Haploids ?
Speed in Line Development
Founder line 1 x Founder line 2
F1Selfing
Inbred line
DH
Inbred line
Haploid Induction & Doubling of Allelic Diversity Materials
Andrew Smelser et al., Plant Introduction Station, Ames
QTL MAPPINGType of
PopulationStrength 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
X
Line 1 Line 2
or
Selfing (F2) DH Induction
Goal: Fixation of target alleles
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
Probability for Fixation of Target Genes
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)
Tuvesson et al. 2007, EU-COST
Other species (Tuvesson et al.): Rye, Oat, Triticale, Potato, Cabbage
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
Colored Seed
Smelser et al. 2009
Partial Fertile
Questions relating to inducer:
- Alternative/improved marker systems- Increased induction rate- Adaptation to US climate / photoperiod- Genetics / biology of induction mechanism
Questions relating to genome doubling:
- Genetic background effect ?- Optimized procedure (with colchicine)- Procedure without colchicine ?- Increased doubling / success rate
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 Lü[email protected]
Collaborators:Candy Gardner, Mike Blanco, Uschi Frei
Costs ?Service On Campus Fee Off Campus Fee
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 ?