last  page PLNT3140 Introductory Cytogenetics
Lecture 22, part 1 of 4
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November 30, 2017


SINGH, R.J. 1993. Plant Cytogenetics. Chapter 4. p.54-60.

SCHULZ-SCHAEFFER, J.1980. Cytogenetics. Plants. Animals. Humans.Springer-Verlag NY. Singh ND . Chapter 15.p.244.

Learning checklist:

1. Be able to list four ways of getting haploid plants.
2. Be able to outline the behavior of chromosomes during meiosis in haploids
3. Be able to outline the method for producing doubled haploid plants
4. Be able to define aneuploidy, trisomy, tetrasomy, and nullisomy
5. Understand the differences between how plants and animals tolerate polyploidy
6. Understand the  mechanisms that result in the formation of chromosome chains in trisomics.
7. Understand how phenotypic ratios are calculated for testcrosses and for the F2 generation of selfed plants
8. Understand why trisomies are usually lethal in humans, and why certain chromosomes can be trisomic.
9. Understand how nullisomic lines of wheat can be used to assign the chromosomal location of a recessive allele.

A. Naturally occurring haploids

Haploidy- is a general term designating individuals or tissues (in mosaics) that have somatic cells with a gametic chromosome number (n). In cases of polyploid series in which there are different species carrying multiples of the basic chromosome set (x) the term haploidy is subdivided into

1. monohaploids(x) - individuals arising from diploid species

2. polyhaploids (2x,3x,4x,etc.) Individuals that can arise from any given polyploid species ( 4x,6x, etc.)

Origin of haploids

Haploids can be produced spontaneously or by induction. Spontaneous haploid production through asexual development as a haploid of an individual that should be diploid was discovered in Drosophila, salamander, newt, frogs, mouse, and chicken. Haploids in animal species are physiologically abnormal and die during embryogenesis.

Examples of spontaneous haploid production are found in a number of plant species including tomatoes, cotton, coffee, beets, barley, flax, coconut, pearl millet, rapeseed asparagus and wheat. In all, there are 36 species of 26 genera and 10 families in which haploidy occurred spontaneously.

B. Methods to generate haploids

Methods of haploid induction in plant species:

1. interspecific and intergeneric hybridization

2. irradiation and chemical treatment

3. selection of twins

4. anther and pollen culture


1. Interspecific or intergeneric hybridization:

In a wide cross between Solanum nigum using pollen from S. luteum,  7 of the 35 S. nigrum progeny were haploid. This mechanism is termed matroclinal pseudogamy in which the male gamete is required to stimulate embryo development. Then the embryo develops directly from the egg without fertilization. This technique is used as a breeding technique in potatoes.
Wheat crossed to a wild species such as Aegilops caudata X Triticum astivum (pollen parent) and then backcrossed to wheat produces a high frequency (53%) of haploids.

Maize has been used as the pollen parent for wheat to produce high frequency of haploids. The maize chromosomes are found in varying numbers in the first cells of the embryo but are then selectively eliminated to leave only the wheat genomic complement. 

In crosses between cultivated barley Hordeum vulgare 2x=14 with wild relative H. bulbosum, H. bulbosum chromosomes are usually lost in the developing embryo. The yield of haploids is relatively low: 11% to a high of 68%. The loss of chromosomes is gradual so that 3-5 days after pollination, 40% of dividing cells are haploid. After 11 days post pollination, 94% are haploid.

Table 4.7. Types and Frequencies of Progeny Obtained from Interspecific Crosses between Hordeum vulgare (V) and H. bulbosum (B)

Genotype and chromosome number
Expected genomic constitution
Cross combination 

(Female X Male)

No. of plants








VVBB Embryo Endosperm
VV x BB 1544 1517
26 1
1V:1B 2V:1B
BB x VV 35 35

1V:1B 1V:2B
VV x BBBB 87

1V:2B 2V:2B

1V:2B 1V:4B

2V:1B 4V:1B

3 2V:2B 4V:2B

2V:2B 2V:4B
Modified from Kasha, 1974.

Many mechanisms have been proposed for chromosome loss, including chromosome fragmentation, degradation of chromatin, lagging chromosomes or bridges, non-congressed chromosomes at metaphase or failure of chromosome migration to the anaphase poles. It may also be related to differences in duration of somatic cell cycles in two parents (asynchrony) as the somatic cycle is longer in H. bulbosum.

2.Irradiation for haploid production.

Plants are pollinated with irradiated pollen which has lost its ability to fertilize. The pollination stimulates the unfertilized eggs to parthenogenic development. Examples are found in various crops including tobacco, wheat, snapdragon, and Oenothera

3.Selection of twins

Selection of haploids can also be achieved by screening for poly-embryonic seed which are seeds with two or more embryos within the same seed coat. The embryos may be haploid or diploid. In Capiscum (pepper), the frequency of twins is controlled by the genotype of the female parent. Through selection, it is possible to raise the percentage of twinning to several percent.

4. Anther and pollen culture

Anther and pollen culture is tissue culture method of production involving culturing of anthers. Haploids develop directly from pollen grains in culture, either through direct formation of embryos from pollen grains or formation of callus and subsequent plant regeneration. This technique requires optimizing culture conditions which vary between species and between genotypes. Induction frequency can be high and the production time to mature plants of three to four months with direct development of homozygous individuals.

C. Meiotic Behaviour of Haploids

Monohaploids have one basic genome (x). Meiosis is very irregular with single homologues present. Chromosomes pair intra-genomically because ancient polyploidization events have left large portions of the genome duplicated. Pairing has been observed in rice, tomato, maize and barley in which synaptinemal complexes appear similar in nature to that found in diploids. Pairing of non-homologous chromosomes in normal diploids is probably evidence that the molecular mechanisms for pairing at zygotene act preferrentially when two chromosomes are nearly identical, as opposed to chromosomes which have diverged over time.

At Metaphase I, the spindle is highly disorganised. At Anaphase I , the distribution of chromosomes to opposite poles is apparently random.

Polyhapaloids originate from autopolyploids or allopolyploids. In autopolyploids, all genomes (x) have the same origin AAAA. In alloploids, basic genomes are of different origin AABB

The classification of autopolyhaploids AA or allopolyhaploids AB can be made on pairing behaviour. Complete absence of pairing is an indication of allopolyhaploids. The occurrence of pairing in polyhaploids is indicative of some chromosomal relationship, homologous or homoeologous

Pairing in haploids is a way of observing ancient duplications. Homoeology describes partial homology during which some chromosome segments pair and some do not pair. Inter-genomic pairing between homolgous or homoeologous chromosomes takes precedence over intragenomic pairing.

Pairing occurs between partially homologous chromosomes of allopolyhaploids. This has been observed in wheat, considered to be an allopolyploid AABBDD. In wheat, the Ph1 (pairing-homeologous) locus on chromosome 5B encodes a gene product which inhibits pairing between homeologous loci. Ph- mutants have increased homeologous pairing. There is a similar genetic control of chromsome pairing in other species such as oats.  

D. Application of doubled-haploids in plant breeding

One of the major problems facing plant breeders is the simple fact that offspring from a cross have a great deal of variability for important traits such as winter hardiness. Tissue culture makes it possible to create haploid plants, whose chromosome numbers are then doubled to produce normal diploids, bearing two identical copies of every gene. Anther tissue containing microspores are placed on tissue culture media, and grown to become haploid embroys. Embryos are transferred to individual plates and develop into plantlets. Plantlets are then transferred into soil. Seedlings are sprayed with colchicine, which causes chromosome doubling.  When plants flower, the germline cells will be diploid. Offspring of doubled haploid plants are therefore more genetically uniform, for all important traits. Homozygosity allows direct selection of recessive characters and simplified gametic ratios. 

Canola (Brassica napus ) embryos regenerating from haploid microspores.
Peijun Zhang, Ph.D. thesis University of Manitoba, 1998
Regenerating haploid plantlet.
Peijun Zhang, Ph.D. thesis University of Manitoba, 1998

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