1. Polyploidy – so many options

2. Impacts of Ploidy Changes
Changes in chromosome number and structure can have major health impacts e.g. trisomy 21
Polyploidy in cultivated and domesticated plants is widespread and of evolutionary and economic importance

3. Polyploidy – Pros and Cons
Advantages
Vigor effects – heterotic boost from divergent parental genomes
Redundancy – masking of recessive alleles
Buffering capacity
Disadvantages
Changes in cell structure & shape – doubling genome content increases cell volume
Problems in cell division – mitosis and meiosis
Changes in gene expression
Epigenetic instability

4. Alternation of Generations
The sporophytic generation may be diploid (2n = 2x) or polyploid (2n = _x)
VAVA
VAVAVBVB
VAVAVBVBVDVD
2n = 2x = 14
30,000 genes
2n = 4x = 28
60,000 genes
2n = 6x = 42
90,000 genes
1 pair homologous chromosomes
0 sets of homoeologous chromosomes AA
2 pairs of homologous chromosomes
2 sets of homoeologous chromosomes
AABB
3 pairs of homologous chromosomes
3 sets of homoeologous chromosomes
AABBDD
A A B B
A A
A A B B D D

5. Euploidy
An organism with an exact multiple of a basic chromosome number (x)
Can be diploid (2x), triploid (3x), tetraploid (4x) …..
Barley in the sporophytic generation is 2n = 14
n = 7 in the gametophtyic generation
The base number (x) = 7 = n for a diploid
Potato in the sporophytic generation is 2n = 48
n = 24 in the gametophytic generation but x = 12
2n=2x=14
2n=4x=48

6. Aneuploidy
Not euploid – more or less chromosomes than a multiple of the basic number
Monosomic – loss of a chromosome, (2n-1)
Trisomic – addition of a chromosome, (2n+1)
Of value in genetic studies

7. Polyploidy More than two basic sets of chromosomes
Autopolyploidy – 3 or more copies of each chromosome in the basic number
Allopolyploidy – 2 or more copies of ancestral genomes giving 4 or more copies of the basic number

8. Polyploid Formation Genome duplication
Failure of spindle fibers in meiosis or mitosis
Must lead to balance in order to achieve euploidy and long-term viability
Imbalanced gametes = aneuploidy

9. Polyploids - Bivalent Pairing for viability
Trivalent + of homologous chromosome pairing in autopolyploids = sterility
Homoeologous pairing in allopolyploids = sterility
If non-bivalent pairing, gametes will not all get the same number of chromosomes
Must have bivalent pairing for fertility: polyploids behaving as diploids

10. Autopolyploids
Three or more homologues for each chromosome in the basic number
Even numbers (4x, 6x etc.) can be fertile
Potato 2n = 4x = 48; Alfalfa 2n = 4x = 32
Pairs of homologs = bivalents and normal meiosis
Odd numbers (3x, 5x, …) = sterile or abnormal
Banana 2n = 3x = 33
The complete chromosome complement cannot form into pairs and normal meiosis is disrupted

11. New Autopolyploids
Can be synthesized by the use of colchicine to double the chromosome complement
Colchicine interferes with spindle formation in cell division
A 2n homozygous cell undergoes replication of each chromosome during S phase of mitosis giving 2 copies of each
No spindle at Anaphase and all can migrate to the same cell to give a homozygous tetraploid

12. New Autopolyploids
Can also create triploids by crossing related tetraploid with a diploid
Newly synthesized autopolyploids generally sterile
Formation of multivalents disrupts meiosis
Advantage in breeding some crops
Seedless watermelon 2n = 3x =33

13. Genetics & Breeding of Autopolyploids
Potentially very complex as up to 4 copies of an allele at each gene can be present
Nulliplex, simplex, duplex, triplex, quadriplex …..
Cross
Nulliplex (N)
Simplex (S)
Duplex (D)
Triplex (T)
Quadriplex (Q)
Nulliplex (aaaa)
All N
Simplex (Aaaa)
1S : 1N
1D : 2S : 1N
Duplex (AAaa)
1D : 4S : 1N
1T:5D:5S:1N
1Q:8T:18D: 8S:1N
Triplex (AAAa)
1D : 1S
1T : 2D: 1S
1Q:5T:5D:1S
1Q : 2T : 1D
Quadriplex (AAAA)
All D
1T : 1S
1Q : 4T : 1D
1Q : 1T
All Q

14. It’s all Bananas
Cultivated bananas derived from diploid species Musa acuminata (A) and Musa balbisiana (B)
Most edibles are triploids with genomes of AAA (desert), AAB (plantains), and ABB (Cooking)
Irregular pairing means bananas are seedless
Good for the consumer but problematic for the breeder and maintainer
Evidence of pairing between homoeologous chromosomes from A and B genomes
90% desert bananas are cv. Cavendish

15. Sequencing to the rescue?
Previous breeding efforts have looked at mutation
Now major effort resulted in sequencing a wild Musa acuminata genome (AA)

16. Seedless Watermelons
An infertile triploid created from 4x and 2x parents
Tetraploid Inbred
AAAA
Diploid Inbred AA x
Triploid F1
AAA
Grow with Fertile Diploid
to stimulate seedless fruit production

17. Seedless Watermelons
The consumer benefits but breeding is more difficult and hence expensive
Development of suitable tetraploids
Selection against sterility and fruit abnormalities
Reduced seed yield for seed company
Grower devotes up to 33% field to 2x pollinator

18. Allopolyploids
An individual with chromosome sets from two or more different but related species
Interspecific hybridization followed by chromosome doubling
Spontaneous (natural forms)
Colchicine (synthesized forms)
Behave like diploids due to bivalent pairing
Homologs within each ancestral species pair even though homoeologous genomes may be collinear

19. Incipient allopolyploids – sterile unless there is doubling
Interspecific hybrids have just one copy of each genome
AA x BB → AB
Haploid number of chromosomes from each species
Gametes get the wrong number of chromosomes and hence infertility
Use colchicine to double the chromosome complement

20. Polyploidy in the Triticeae
Sporophytic generation
Gametophytic generation
Ploidy Level
Genome Formula
2n = 14
n = 7
2x (diploid) e.g. Emmer wheat
2n = 2x = 14
2n = 28
n = 14
4x (tetraploid) e.g.
Durum wheat
2n = 4x = 28
2n = 42
n = 21
6x (hexaploid) e.g. Bread wheat
2n =6x = 42

21. The Evolution of Bread Wheat (and barley)
Hordeum spontaneum
Wild barley 2n = 2x =14
AA=BB=DD 2n=2x=14
Hordeum vulgare
Cultivated barley 2n = 2x =14

22. The Bread Wheat Genome

23. For more information see
Wheat Pairing
Ph1 locus on 5B affects pairing in wheat
Promotes homologous pairing
Blocks homoeologous pairing
Gene has been cloned
For more information see

24. Triticale an allo-hexa/octaploid
Wheat (durum or bread) Rye hybrid
Bread Wheat
AABBDD
Rye RR x
Infertile F1
ABDR
Fertile F1
AABBDDRR

25. Brassicas – The Triangle of U (Woo Jang-choon = Nagahara U)

26. Haploids Single basic set of chromosomes
Maize – n=10; bread wheat – n=21; barley – n=7
Haploid plants can be nurtured to grow
Only have the basic chromosome content (n) so are infertile – meiotic irregularities

27. Doubled Haploids
Doubling the haploid chromosome content gives two exact copies
No heterozygotes – “instant inbred lines”
Sample pollen or egg cells from F1 plants
A random sample of all the possible products of the first round of segregation from meiosis
Shorten the breeding cycle
Immortal genetic populations for research
Can make doubled haploids at any stage in the selfing process (e.g. F1, F2, F3)

28. Doubled Haploids Production of ‘Instant’ Inbreds
Shortens Breeding Cycle
Makes Selection More Effective
Can make Pure Seed Production Easier
How?
Pollination by alien species
Anther/Microspore Culture
Haploid inducing genes

29. Doubled Haploidy Time Line
1921: Natural production of haploids in Datura stramonium observed. Followed by Nicotiana tabacum (1924)
1952: Doubled haploid, inbred maize lines produced.
Selected parthenogenic haploids and chromosome doubling
1964: Haploid plants from Datura innoxia by anther culture
1970: Haploid production in barley via wide crossing
1978/79: First doubled haploid cultivar: “Mingo” barley
Currently: Routine technique in breeding many cereal and vegetable crops

30. Events in Androgenesis
maturation
bi-cellular
pollen
mature pollen
male gametophyte
stress
uni-cellular
microspore:
embryogenesis
embryo, sporophyte
embryogenic microspore
cell with restricted
developmental
potential
Diverting the developmental pathway of the microspore towards embryos rather than pollen
A. Touraev: Vienna Biocenter, University of Vienna, Dr. Bohr-Gasse 9, A-1030 Vienna, Austria
totipotent cell

31. Androgenesis Induction
Reprogramming of microspores
towards
sporophytic development
Heat shock
Sucrose and
nitrogen starvation
Cold stress
Colchicine treatment
Re-programming of the microspore.
A. Touraev: Vienna Biocenter, University of Vienna, Dr. Bohr-Gasse 9, A-1030 Vienna, Austria
Gamma irradiation
Ethanol pH
Osmotic stress
Separate or in combination

32. Hordeum bulbosum wide crosses
Produce F1 from Desired Cross
Emasculate 2-3 days before pollen shed
Ensure plentiful supply of pollen from wild relative
Dust alien pollen onto open emasculated flowers
Apply hormonal spray to pollinated spike (can repeat 2-3 days later)
Bag pollinated spike and leave for days
The Hordeum bulbosum method to produce doubled haploid barley.
P. Devaux: Florimond Desprez, Biotechnology Laboratory, P.O. Box 41, Cappelle en Pevel,e, France.

33. Hordeum bulbosum wide crosses
Rescue developing embryos from spike pollinated with alien pollen
Grow on in special rooting medium
Once plants established, trim roots and treat with colchicine
The Hordeum bulbosum method to produce doubled haploid barley.
P. Devaux: Florimond Desprez, Biotechnology Laboratory, P.O. Box 41, Cappelle en Pevel,e, France.
Grow out plants and harvest seed from fertile plants

34. Anther Culture Healthy Donor Plants Harvest spikes
Apply stress conditions
Plate out anthers on induction medium
Sub-culture steps
Spontaneous doubling
Transfer to greenhouse
Field

35. Numbers of DH Cultivars
Species
Numbers
Method
Rice
>100
Anther culture
Barley
Anther, micospore culture & wide crossing
Rapeseed
>50
Microspore culture, spontaneous DH lines
Wheat
>20
Anther culture, wide crossing
Pepper
>10
F1 from DH parent(s)
Asparagus
Female x DH supermale
Tobacco
Microspore culture, anther culture
Cultivars derived through doubled haploidy
B.P. Forster: Scottish Crop Research Institute, Invergowrie, Dundee, DD2 5DA, UK.
Also: mustard, eggplant, melon, triticale

36. Doubled haploids & inbred line development in maize
Hybrid maize (corn) is a major crop worldwide
Hybrids derived from intermating inbred lines
Inbred line development key to hybrid breeding
Accelerate inbred line development means hybrid development also accelerated
In vitro production of doubled haploids
Anther or microspore culture
In vivo production of doubled haploids
Haploid inducer lines either as male or female
Induction at >1% haploid lines; morphological marker for identification
Possibly arise through defective sperm cell enabling fertilization but chromosomes eliminated