POLYPLOIDY BREEDING

            Polyploidy Breeding



Introduction:

 Plant breeding is being playing significant role for the improvement of the characteristics of the plants. That is required for the development of the new variety successfully and economically. Plant breeding is being used for getting the desirable characters from different sources. There are different types of breeding techniques have been developed for the crop improvement. Hybridization is being utilizing for creating genetic variation. Stebbins (1958) defined hybridization as the “crossing between individuals belonging to separate populations which possess different adaptive norms”. Beside Mendelian variation, comprising of mutation coupled with or without hybridization between mutant forms, polyploidy also contributes to the evoluation of crop species. Polyploidy, a prime facilitator of speciation and evolution in plants and to a lesser extent in animals is associated with intra and inter-specific hybridization. (Levin, 1983) Polyploidy is an intriguing phenomenon in plants that has provided an important pathway for evolution and speciation. Although the first polyploid was discovered over a century ago, the genetic and evolutionary implications of polyploidy are still being elucidated (Bennett, 2004; Soltis et al., 2003). Polyploidy provides the opportunity for selection to sculpt a variety of new gene functions, traits, and lineages. Many important model systems, both agricultural and wild, have been used to study the consequences of recent polyploidization (e.g., cotton [Gossypium hirsutum], tobacco [Nicotiana tabacum], wheat [Triticum aestivum], canola [Brassica napus], soybean [Glycine max], potato [Solanum tuberosum], sugarcane [Saccharum officinarum], cordgrass (Spartina anglica), and dandelion [Taraxacum officinale]) (Wendel, 2000; Osborn et al., 2003; Tate et al., 2005). the most extensive evidence for ancient polyploidy comes from analysis of the complete sequences of both rice (Oryza sativa; Yu et al., 2005) and Arabidopsis.

Polyploidy:

An Organism or individual having more than two basic or monoploid sets of chromosomes called polyploid and such condition is known as polyploidy. It has been also defined as the possession of three or more complete sets of chromosomes and has been an important feature of chromosome evolution in many eukaryotic taxa including plants, yeasts, insects, amphibians, reptiles, fishes and even the mammalian genome (Ramsey and Schemske, 1998). According to Grant (1971, 1981), the phenomena of polyploidy was discovered during the exploratory phase of plant cytogenetics in the early years of the twentieth century. Winkler (1916) introduced the term polyploidy, and Winge (1917) proposed that polyploidy occurred by somatic doubling in species hybrids. Polyploidy has been regarded as a major force in evolution and speciation (Schultz, 1980; Soltis, 1995). There are several examples of polyploid animals (Mable, 2004), including mammals (Gallardo et al., 1999), fishes (Le Comber and Smith, 2004), and frogs (Ptacek et al., 1994), and About 30-35% of the angiosperm species are polyploidy (Stebbind, 1958). There are about one third species of flowering plants, 70% of wild grasses family are polyploids. It is rare in animals because it’s lethal effect. But polyploidy found in those animals which are developed parthenogenetically like aphids. Polyploidy is rather common in some families like Rubiaceae, Compositae, Iridaceae, Gramineae etc. It is uncommon but present in other families such as Caesalpinaceae, Passifloraceae and Fagaceae (Grant, 1981). The highest percentage is found in perennial herbs and a smaller proportion is found in annuals and woody plants (Stebbins, 1950; Stebbins, 1971).

Polyploid is under Euploids belongs to the group of Heteroploid. When the number of genomes or copies of a single genome more or less than two is called euploid, and the situation is called euploidy. And the individuals carrying chromosome numbers other than the diploid (2x, and not 2n) number are known as heteroploid, and the situation is known as heteroploidy.

The phenomenon of polyploidy gained much of what it is today during the early part of the twentieth century. One of the early examples of a natural polyploid was one of De Vries’s original mutations of Oenothera lamarckiana (mutat. gigas)( Ramsey and Schemske, 1998). The first example of an artificial polyploid was by Winkler (1916) who in fact introduced the term polyploidy (Grant, 1981; Ramsey and Schemske, 1998 ). Winkler was working on vegetative grafts and chimeras of Solanum nigrum and found that callus regenerating from cut surfaces of stem explants were teratploid (Grant, 1981; Ramsey and Schemske, 1998). Digby(1912) had discovered the occurrence of a fertile type Primula kewensis from a sterile inter-specific hybrid through chromosome doubling but failed to realize its significance in the context of polyploidy(Stebbins, 1971). Though unaware of the ‘Primula type” fertile hybrid, Winge (1917), from his studies on the chromosomal counts of Chenopodium and Chrysanthemum found that chromosome numbers of related species were multiples of some common basic number; he subsequently proposed a hypothesis that chromosome doubling in sterile inter-specific hybrids is a means of converting them into fertile offsprings (Swanson et al, 1981; Hieter and Griffith, 1999). This was subsequently verified by various workers in artificial inter-specific hybridizations of Nicotiana, Raphanobrassica and Gaeleopsis (Grant, 1981).Finally the colchicine method of chromosome doubling was developed by Blakeslee and Avery (1937) and became an important tool for the experimental study of polyploidy ( Song et al, 1995). many of our crop species, including wheat, maize, sugar cane, coffee, cotton and tobacco, are polyploid, either through intentional hybridization and selective breeding (e.g. some blueberry cultivars) or as as a result of a more ancient polyploidization event(e.g. maize)( Ramsey and Schemske, 2002). The importance of polyploidy in such diverse fields such as cytogenetics, physiology, breeding, cytotaxonomy and biogeography in conjunction with new possibilities put forth by various molecular techniques has all spurred a resurgence of interest in issues of origin and establishment of lineages(Mable, 2003). Polyploidy seems to be favored in long lived/perennial plants possessing various vegetative means of propagation (eg: Fragia, Rubus, Artemisia, Potamogeton etc.) and in those with frequent occurrences of natural inter-specific hybridizations (Hilu, 1993).

Polyploidy Breeding:

Polyploidy breeding refers to induced chromosome manipulation. Success is wholly dependent on the control of chromosome pairing and recombination in polyploids and their hybrid derivatives. Breeding strategies for transferring genes across ploidy levels depend on their origin.

Types of Polyploidy: 1. Autopoly ploidy

 a. Autotriploidy (3x)

 b. Autotetraploidy (4x)

c. Autopentaploidy (5x)

 d. Autohexaploidy (6x)

2. Allopolyploidy

a. Allotetraploidy (2x1+2x2)

 b. Allohexaploidy (2x1+2x2 + 2x3)

 c. Allooctaploidy (2x1+2x2 + 2x3 + 2x4)

Autopolyploids:

Polyploids which originated by doubling of the chromosome number of a diploid species, or a hybrid between races of the same species, resulting in two pairs of chromosomes is called autopolyploid, and the condition is referred to as autopolyploidy. In mode of origin,autopolyploids arise within single populations or between ecotypes of a single species (M¨untzing, 1936; Darlington, 1937; Burnham, 1962; Gottschalk, 1978) and in cytological criteria, autopolyploids will exhibit multivalent configurations, nonpreferential pairing at metaphase, and multisomic inheritance (Stebbins,1980; Jackson 1982; Jackson & Jackson 1996). 1. Autotriploids: Present of three full sets of chromosomes of the same species. They are highly sterile due to defective gamete formation. It is useful for asexually propagated plants like banana, sugarcane, apple, watermelon, sugar beet etc.

2. Autotetraploids: Present of four full set of chromosomes of the same species. They are very stable and fertile as pairing partners are available during meiosis. They are usually larger and more vigous than the diploid species. It is applicable in rye, grasps, alfalfa, groundnut, potato, coffee etc.

Origin of autopolyploids: Autopolyploids may be originated by any one of the following several ways:

1.       Spontaneous: Chromosomes become double occasionally in somatic tissues and also form unreduced gametes in low frequencies due to certain genes, e.g., genes causing complete asynapsis or desynapsis. Spontaneous somatic chromosome doubling is a rare event (Lewis, 1980) and the only well documented instance of the same was in case of tetraploid Primula kewensis which arose by somatic doubling in certain flowering branches of a diploid hybrid.

2.  Production of adventitious buds: Callus developed at the cut end of stem due to decapitation may have some polyploidy cells which are commonly found in Solanaceae.

3. Physical agents: Polyploidy may be occurred due to heat or cold treatments, centrifugation and X-ray or gamma ray irradiation in low frequencies. Development of tetraplod branches in Datura due to cold treatment. 2-3% tetraploid progey may produce due to heat treatment of 38-45ºC on the maize plant or ears. Successful application of heat treatment on barley, wheat, rye and some other crops have been found.

4. Regeneration in vitro: Some of the plants regenerated from callus and suspension culture may be polyploids as foun in the Nicotiana, Dautura, Oryza sativa etc.

5. Colchicine treatment: It is the most effective and the widely used treatment for polyploidy production. Colchicine is an alkaloid, which is found in the seeds and bulbs of autumn crocus (Colchicum autumnale). It can be used in aqueous solutions, in paste with lanolin, in glycerine or in agar. It’s low concentrations are applied to the lethal buds or growing tips of the desired plants. Colchicine acts as spindle suppressors at mitosis and results in cells with double number of chromosomes per cell. Some examples of colchicines treatments are as-

a. Seed treatment: Seeds are treated with colchicines of 0.001-1% for 1-10 days.

b. Seedling treatment: Shoots of young seedlings are treated with colchicines for 3-24 hours.

c. Growing shoot apices: These are treated with 0.1-1.0% colchicines for once or twice daily for a few days.

d. Woody plants: 1% colchicines is generally used for application on shoot buds.

6. Other chemical agents: Use of some other chemicals i.e., acenaphthene, 8- hydroxyquinoline, nitrous oxide etc., which have polyploidizing effect.

Effect of Autopolyploidy on Plants:

 Autopolyploidy leads to production of larger cell size than the diploid which is termed as ‘giggas’ properties (Briggs and Knowles, 1967). The leaves of polyploidy are generally ticker and larger than diploid form. Vitamin A activity in tetraploid maize is increased as compared to the diploid (Randolf and Hand, 1940) It is about 40% more than diploid. Similarly, the vitamin C content of vegetables and fruits has been known to increase following chromosome doubling. The nicotine content of tetraploid tobacco is about 18- 33% higher than in diploid species. Growth rate of polyploidy is relatively slower than the diploid (Briggs and Knowles, 1967). Self-incompatibility systems observed in diploid species are often weakened or may even disappear in polyploidy. Reduced fertility due to meiotic irregularities may be found. Larger pollen grain, presence of larger and thicker leaves larger flowers and fruits, in many cases, increased vigour and vegetative growth, containing lower dry matter than diploid in polyploids. In some cases reduction in fertility as compared to their counterpart may be as high as 80-95%.

Genetics of Autopolyploids:

1. Meiosis in Autopolyploids:

 a. Double reduction in autotetraploids:

In a locus autopolyploidy segregation may be random chromosome segregation (RChS) or random Chromatid Segregation (RCdS). Under RChS, the two sister chromatid of a chromosome never assort into the same gamete. Under RCdS, The two sister chromatid may segregate into the same gamete. This Phenomenon is known as “double reduction”, is a specific property of autopolyploids.

At anaphase I, migration of sister chromatid i) to the same pole to give reductional separation. (Type I in Figure 1.2), or ii) to different poles to give equational separation ( Type II & III in Figure 1.2). This latter separation depends on the occurance of a crossover between the locus under consideration and the centromere. With no crossover between the centromere and the locus the first division is necessarily reductional and the second equational, so that double reduction can not occur (Type I). If there is crossover, sister chromatids are associated with different centromeres, though the occurance of a double reduction still depends on the poles to which the two sister chromatids migrate. If they migrate to different poles, the separation at anaphase II is still equational, i.e., there is no double reduction (Type II). If they migrate to the same pole at anaphase I, the second division can be reductional and lead to double reduction. When chromatids segregate randomly, double reduction is expected to occur in half the cases (Type III). Such a meiosis is sometimes also called “pseudo – equational” (Mather, 1935. the consequences of this phenomenon is that two copies of the same gene i.e., identical genes, can be in the same gamete. This means that double reduction generates inbreeding even in the absence of consanguineous mating.

b. Double reduction in other autopolyploids:

Duble reduction defined as a phenomenon which leads to the presence in a gamete of copies at the meiosis of the same gene may affect all allopolyploids. For autohexaploids, the extension is straightforward, as shown by Fisher (1949). Extension to higher ploidy level is more complex because the double reduction may affect two genes within a gamete for octosomic or more. For octosomics, with only quadrivalents randomly formed at the meiosis, analogous to that defined for autotetraploid, is sufficient. Then form a genotype abcdefgh only three kinds of gametes are possible abcd, aabc, and aabb.

c. Gametic Association of Alleles:

A consequences of meiosis in autoploid genetics is that the association of genes in gametes is not wholly random. For example, a tetraploid AAaa produces gametes in the proportion of 1/6 AA : 2/3Aa : 1/6aa (assuming RChS) while under random association, the proportion would be 1/4AA : 1/2 Aa : 1/4aa. This is due to the fact that an individual produces gamete that are either parental or recombinant. Genes associated in a gametes at the origin of one individual tend to remain associated. The relationship between alleles at a single locus is similar to that between non-homologous genes for diploids (Gallais, 1974)

The Production of Unreduced Gametes In diploid:

 The production of unreduced gametes, that is gametes with the somatic chromosome number, is involved in the origin of natural polyploidy plants.

Cytology in Autoploids

 Autoploid contain more than two homologous chromosomes. Consequently, instead of forming bivalents during meiosis as diploid, there are also multivalent .

For example, autopoids have mostly trivalents but some bivalents and univalents are also present. Tetraploids have qudrivalents or bivalents as well as some trivalents and uunivalents. These meiotic abnormalities are implicated in sterility to some extent, more so in triploids. The microspores and megaspores with x or 2x genomes are usually viable. The amount and nature of chromosome pairing directly impacts the breeding behavior of autoploids. Autoploids are induced artificially by chromosome doubling using colchicines. The tendency for the coupled set of chromosomes from one parent to pair independently of the doubled set of chromosome of another parent is called preferential or selective pairing. The concept of preferential pairing is applied in the modern breeding of polyploids whereby alloploids are stabilized and make reliable as diploids, a process called diplodization.

Use of Autopolyploid in Plant Breeding:

 Following three factors should be taken in consideration as a guide for production and utilization of the autopolyploid in plant breeding:

 i) Autopolyploid manifests greater vegetative growth but reduced seed production. This implies that autopolyploid induction would be more useful for vegetative parts of the plants, such as forage, or root, but not the seed.

ii) Autopolyploid produced from diploids with lower chromosome number have been relatively more successful.

iii) Performance of a diploid species in a given environment may not be correct indication regarding the performance of corresponding autotetraploid produced from it. Cross pollinated crop may be more successful as parents for autopolyploid induction than self pollinated species (Poelman, 1987). The reason is obvious. Because cross pollination greater genetic variability will be generated by cross pollinated plant species and consequently a balanced polyploidy genotype may emerge. Successful autopolyploid program were developed in few crops. A few examples are presented below:

a) Triploid Sugar Beets: The roots of triploid sugar beet are larger in size than diploids, but they maintain the sugar content of diploids and hence yield more sugar per unit area. Triploids are highly sterile and do not produce seed. Triploid sugar beets are produced by interplanting diploid anf tetraploid. Tetraploid is used as seed parent and triploid seed is therefore harvested from tetraploid. Autotetraploid beets have smaller roots as compare to diploid. Triploid can be produced when tetraploid (individuals with 4 genomes in somatic cells) is crossed with diploid or when something goes wrong in meiosis and unreduced gametes are produced (with 2n chromosomes) which unite with gametes carrying n chromosomes (haploid chromosome number). Duriong the Zygotene and pachytene stages of meiotic cell division three homologous chromosomes (trivalent) synapse but in such a way that a given position only homologous are together. At anaphase I, two homologues may go to one pole and one to another. But considering all the chromosomes within a cell, the 4 meiotic products may contain from n to 2n chromosomes, with all integral values between them. Gametes containing either n or 2n chromosomes may be functional, but the gametes that are deficient for either of the chromosomes or have additional chromosomes are sterile.

b) Triploid Watermelons: Seedlessness would be advantageous in watermelons. Diploid watermelons (Citrullus lanatus) have 22 chromosomes per somatic cell and are fully fertile, and produce large number of seeds per fruit. Natural Parthenogenesis is not known for watermelons. Hence Kihara’s (1951) technique of producing seedless, triploid watermelons can be utilized. The autotetraploid lines (produced by  doubling the chromosome number of a diploid) are planted alternately with diploids in isolation. Tetraploids are employed as seed parent. The triploid seed produced on tetraploids is viable. However, when diploid is used as female, the program of triploid seed production is unsuccessful. Because of meiotic irregularities, triploids produce no true seed but rudimentary structures similar to cucumber seeds (small and white) are formed in triploid fruits. For raising a successful commercial crop of seedless triploid watermelon, it is essential to interplant diploid variety because fruit setting on triploids depends upon stimulus provided by the pollen. Main difficulty with triploid watermelon seed production is maintenance of tetraploid seed parent. Moreover, the triploid seedless fruits are irregular in shape and tend to be hollow. Germination of triploid seed is difficult. Production of triploid seed reqiers hand pollination of tetraploid flowers and hence cost of triploid seed is prohibitive. Because the gametes produced by triploid plant are largely unbalanced with respect to chromosome number, triploid are inviable. If, however, triploid plants could be vegetatively propagated, viz, by cuttings, they could be advantageous.

c) Rye (Secale cereale): Among grain crops, rye is the only crop to be successfully developed as an autotetraploid. Tetraploid rye has larger kernels, superior ability to emerge under adverse condions and higher protein content. It is grown in Sweden and Germany.

d) Tetraploid grapes: Tetraploid grapes has been developed in California, USA which have larger fruits and fewer seeds per berry than diploids.

e) Forage crops: Swedish tetraploid strains of alike and red clover have given higher hay yieds than corresponding diploids. Pusa Giant berseem (a variety of Egyptian clover) was released in India for higher fodder yield.

f) Ornamentals: Induced autopolyploidy has been most successful in ornamentals, for example snapdragons. In such crops novelty itself is a virtue. Autopolyploids may have bigger flower size, longer blooming period and relatively longer lasting flowers.

g) Alfalfa: Tetraploid varieties of alfalfa are better than diploid in yield and have better recovery after grazing.

h) Tetraploid Banana (Musa sapientum): Autotetraploidare inferior to triploids in that they have weaker leaves and increased fertility. But they offer the only available chance of adding disease resistance to commercially successful varieties. In banana, autotetraploids are produced by chance fertilization of an unreduced triploid egg (AAA) by a haploid pollen from a disease resistance diploid parent.

i) Maize: The tetraploid maize has 43% more carotenoid pigment and vitamin A activity than the diploid.

j) Teraploid potato: Crosses between diploid parents with variable expressivity of 2n gametes generally produce tetraploid and diploid offspring (Ortiz and Peloquin, 1991b)

k) Tetraploid turnip: Vegetative growth of tetraploid turnip is much greater than diploid and commercially available in Europe (Poehlman and Sleper, 2006).

Limitation of Autopolyploid:

1. The larger size of autopolyploids is generally accompanied with higher water content. As a result, autopolyploids of the crop species grown for vegetative parts do not always produce more dry matter than the respective diploids. For example, tetraploid turnip (Brassica rapa) and cabbage (Brassica oleracea) outyield the diploids in fresh weight, but are comparable, or even inferior, to them in terms of dry matter production.

2. In crop species grown for seed, autopolyploids show high sterility accompanied with poor seed set.

 3. Fertility in autotetraploid can be increased by hybridization and selection at the tetraploid level. But due to the complex segregation in autotetraploids, progress under selection is slow.

 4. Triploid can not be maintained except through clonal propagation.

5. New polyploids (raw polyploids) are always characterized by a few or more undesirable features, e.g., poor strength of stem in grapes, irregular fruit size in watermelon, etc.

6. Effect of autopolyploidy can not be predicted.

Allopolyploids: Polyploids which have genomes from two or more species are known as allopolyploids or alloploids or hybrid polyploids or bispecies or multispecies polyploids and such condition is referred to as allopolyploidy. In mode of origin, allopolyploids are derived from interspecific hybrids (M¨untzing, 1936; Darlington, 1937; Burnham, 1962; Gottschalk, 1978) and in cytological criteria, allopolyploids are expected to display bivalent pairing, lack of allosyndesis anddisomic inheritance (Stebbins, 1980; Jackson, 1982; Jackson and Jackson, 1996). Allopolyploids with genomes of two diploid species are allotetraploids or amphidiploids. It has been playing significant role in crop evolution than autopolyploidy as it is found in about 50% of crop plants. Origin of Alloploids: Nowadays, alloploids are mostly produced by chromosome doubling in F1 hybrids between two distinct species (distant hybrids) belonging to the same genus or to different genera. Chromosome doubling might have occurred in somatic tissues due to an irregular mitotic cell division leading to the formation of allopolyploid sectors either in the apical meristem or in axillary buds; the latter would produce allopolyploid branches. Sexual progey of this branches would be allopolyploids. On the other hand, irregular meiosis may produce unreduced gametes. They may unite to produce allopolyploid progeny. Experimental allopolyploids production is achieved by chromosome doubling in distant hybrids with the help of colchicines or some other agent. The allopolyploids produce by man by using two distant species of plants are termed as synthetic allopolyploids. However, allopolyploids productions involve two steps as, i) production of F1 distant hybrids, and ii) chromosome doubling .

Effect of Allopolyploid on plant:

It is very difficult of predicting the combine parental characters that would appear in an allopolyploid. For example, during the production of Raphanobrassica the aim was to synthesize a crop species that would combine the root of Raphanus sativus with leaves of Brassica oleracea. But the Raphanobrassica gave the result totally opposite of the aim. Alternatively, Triticale has combined the desirable features of two parental species i.e., the hardiness of rye (Secale cereale) and the yielding ability of wheat (Triticum turgidum) (Fig. 1.4). Generally, Allopolyploids are highly vigorous than diploids with some exceptional. Though their adaptability may differ from their parents, the evolution of allopolyplody in nature has been favored by the availability of new ecological areas for the establishment of allopolyploids .Most of the allopolyploids are apomictic which are mostly found in grasses, e.g., Poa, and also in many other plant groups, e.g., Taraxacum, Parthenium, Rubus, Fertillica, Tulipa, Solanum, etc.

Genetics of Allopolyploids:

Allopolyploid speciation often results in more individual genetic variation than in a diploid species because different parental alleles combine to form heterozygous isozymic patterns in the allopolyploid (e.g., Ranker et al., 1989; Sun, 1996). For example, Roose and Gottlieb (1976) found ;30–40% ‘‘fixed’’ heterozygosity across loci within the allopolyploids of Tragopogon. Increased heterozygosity, the generation of novel heteromeric enzymes, and the formation of new gene combinations may be critical in the successful establishment of polyploids (Thompson and Lumaret, 1992).

Alloploids arise from the combination and subsequent doubling of different genomes, a cytological event is called alloploidy. The genomes that are combined differ in degrees of homology, some being close enough to pair with each other, whereas others are too divergent to pair. Sometimes, only segments of the chromosomes of the component genomes are different, a condition that is called segmental alloploidy. Some of the chromosome of one genome may share a function in common with some chromosomes in a different genome. Chromosomes from two genomes are said to be homeologous when they are similar but not homologous (identical).

Most alloploids have evolved certain genetic systems that pairing occurs between chromosomes of the same genome. A classic example occurs in wheat (2n = 6x = 42) in which a gene on chromosome 5B, designated Ph, enforces this diploid-like pairing within genomes of the alloploid. When this gene is absent, pairing between homeologous chromosomes, as well as corresponding chromosomes of the three genomes, occurs, resulting in the formation of multivalents at meiosis I.

Alloploids exhibit a variety of meiotic features. Sometimes chromosomes pair as bivalent and thereby produce disomic ratios. Where the component genomes have genes in common, duplicate factor ratios will emerge from meiosis, an event that sometimes is an indication of alloploid origin of the species. Whereas significant duplications of genetic material have been found in wheat, the genomes of upland cotton have little duplication. Tetrasomic ratios are expected for some loci where multivalent associations are found in allotetraploids.

Role of Allopolyploid in Evolution:

A.      Natural Alloploids: Some important natural allopolyploid crops are wheat, cotton, tobacco, mustard, oat etc. Interspecific crossing followed by chromosome doubling in nature have resulted in the origin of allopolyploids. The origins of some natural allopolyploid crops are given bellow:

1.        Bread Wheat (Triticum aestivum):

volutionary history of wheat has been the most extensively investigated. Identity of the diploid species contributing the 3 genomes (A, B and C genomes) of Triticum aestivum has been investigated by many workers more notably by Sears, Kihara and others. It is generally accepted that the A genome present in diploid wheat is the same to those present in tetraploid and hexaploid wheat. Further, the B genome of tetraploid emmer wheats is similar to that found in hexaploid wheat. This is evident from chromosome pairing in crosses among diploid, tetraploid and hexaploid wheat. Hybrids between diploid and tetraploid wheats show about 7II and 7I, while those between tetraploid and hexaploid wheats show about 14II and 7I. It is believed that A genome of wheat has come from Triticum aestivum (2n = 14), D genome from Triticum tauschi (2n = 14) and B genome from unknown source probably from an extinct species (2n = 14).

2.       Tobacco (Nicotiana tabacum):

The genus Nicotiana is a member of the family Solanaceae and comprises 76 currently recognized, naturally occurring species that are subdivided into 13 sections (Knapp et al. 2004). Nicotiana tabacum (n =24) is a classic amphidiploid arisen from the hybridization between Nicotiana sylvestris and Nicotiana tomentosa; both the species are diploids with n = 12. There is a very strong evidence of Nicotiana sylvestris as the maternal parent and the donor of the “S” genome (Bland et al., 1985; Olmstead and Palmer, 1991; Aoki and Ito, 2000; Yukawa et al., 2006). It is highly likely that the “T” genome was contributed by a member of section Tomentosae (Likely Nicotiana tomentosiformis, Nicotiana otophora, or an introgressive hybrid between the two) (Kenton et al., 1993; Riechers and Timko, 1999; Lim et al., 2000b; Kitamura et al., 2001; Ren and Timko 2001).

3.       Cotton:

 Diploid species of the genus Gossypium are all n = 13, and fall into 7 different "genome types," designated A-G based on chromosome pairing relationships (Beasley, 1942; Endrizz, et al. 1984). A total of 5 tetraploid ( n = 2x = 26) species are recognized. All tetraploid species exhibit disomic chromosome pairing (Kimber, 1961). Chromosome pairing in interspecific crosses between diploid and tetraploid cottons suggests that tetraploids contain two distinct genomes, which resemble the extant A genome of G. herbaceum ( n = 13) and D genome of G. raimondii (n = 13), respectively. The A and D genome species diverged from a common ancestor about 6-1 1 million years ago (Wendeil, 1989). The putative A X D polyploidization event occurred in the New World, about 1.1- 1.9 million years ago, and required transoceanic migration of the maternal A genome ancestor (Wendeil, 19 89, Wendeil and Albert, 1992), which is indigenous to the Old World (Fryxel,1979). Polyploidization was followed by radiation and divergence, with distinct n = 26 AD genome species now indigenous to Central America (G. hirsutum), South America ( G. barbadense, G. mustelinum), the Hawaiian Islands (G. tomentosum), and the Galapagos Islands ( G. danuinii) (Fryxelp, 1979). The tetraploid American cotton (Gossypium hirsutumI, 2n = 52) is believed to be an amphidiploid between Gossypium Africana and Gossypium raimondii. Both these species are diploid with 2n = 26. The chromosomes of Gossypium Africana are larger than Gossypium rainondii.

4.       Oat (Avena sativa):

 It is a self-pollinated allohexaploid with a basic chromosome number of n =3x= 21 that consists of three basic genomes (A, C, and D)(Rajhathy and Thomas, 1974) derived from a cross between A. Barbara (tetraploid, n= 14) and A. strigosa (diploid, n=7).

5.       Amphidiploid Brassica Species:

The origin of amphidiploiid Brassica species is presented in Fig 1.6 based on famous U’s Triangle proposed by N.U. in 1935. As per of this Scheme, Brassica juncea (2 = 18) is an amphidiploid from an interspecific cross between Brassica nigra (n = 8) and Brassica campestris (n = 10), whereas Brassica napus (n = 19) is an amphidiploid from an interspecific cross between Brassica oleracea (n = 9) and Brassica campestris; and Brassica carinata (n = 17) is a result of interspecific cross between Brassica nigra (n = 8) and Brassica oleracea (n = 9).

Artificial Allopolyploid:

 Artificial or synthetic allopolyploids have been synthesized in some crops with two main objectives, viz. 1) either to study the origin of naturally available alloploids or 2) to explore the possibilities of creating new species. Some example of artificial alloploids are given bellow:

1. Raphanobrassica: This a classical example of artificially synthesizesd alloploid. This was developed between radish (Raphanus sativus, n = 9) and cabbage (Brassica oleracea, n = 9) by Russian geneticist Karpechenko in 1928. He wanted to develop a fertile hybrid between these two species with roots of radish and leaves of cabbage. But he got a fertile amphidiploid (4n = 36) by spontaneous chromosome doubling which unfortunately had roots of cabbage and leaves of radish. Thus it was of no use.

2. Tobacco: Clausen and Goodspeed synthesized a new hexaploid species of tobacco (Nicotiana) from a cross between Nicotiana tabacum (2n = 48) and Nicotiana glutinosa (2n = 24). The F1 was sterile with 2n = 36, which was made fertile by doubling of chromosome through colchicine treatment. The new species is known as Nicotiana digluta.

3. Triticale: Triticale is a new crop species which has been synthesized from a cross between wheat (Triticum aestivum) and rye (Secale cereale, n = 9). Some triticales are developed from cross between tetraploid wheat (Triticum turgidum) and rye and some from cross between heaxaploid wheat (Triticum aestvum) and rye. The F1 was sterile, which was made fertile by colchicines treatment. Triticales produced using tetraploid and hexaploid wheat are hexaploid and octaploid, respectively. Triticale id now commonly grown in Canada, Mexico, Hungary and some other countries.

4. Wheat: Triticum aestivum (formerly Triticum spelta), is a hexaploid wheat which was artificially synthesized in 1946 by E. S. McFadden and E. R. Sears and also by H. Kihara. They crossed an emmer wheat, Triticum turgidum (formerly Triticum dicoccum) (tetraploid: 2n= 28) with goat grass, Triticum tauschi (formerly Aegilops squarrosa) (diploid; 2n= 14) and doubled the chromosome number in the F1 hybrids. This artificially synthesized hexaploid wheat was found to be similar to the primitive wheat Triticum aestivum. When the synthesized hexaploid wheat was crossed with naturally occurring Triticum aestivum, the F1 hybrid was completely fertile; this suggested that hexaploid wheat must have originated in the past due to natural hybridization between tetraploid wheat and goat grass followed by subsequent chromosome doubling.

5. Cotton: The American cotton (Gossypium hirsutum) was synthesized from a cross between Gossypium tabacum and Gossypium raimondii. Both these species are diploid with 2n = 26. the former is Old World cultivated diploid and latter, New World wild diploid.

6. Brassica species: The synthetic allopolyploids can be produced as like as the natural scheme. The hybrids between the synthetic and natural amphidiploids are reasonably fertile.

7. Holcus lanatus (2n = 2x = 14) X H. mollis (2n= 4x = 28) 8. Triploid hybrid

(2n = 21) that backcrosses to tetraploid H. mollis to form a pentaploid H. mollis (2n = 5x = 35). Jones (1958) used chromosome size and presence of satellites to identify species, i.e. the karyotype.

Spartina (cordgrass) in Europe: The European native species Spartina

maritima (2n = 2x = 60) came into contact in the 1800s with the introduced North American species S. alterniflora (2n = 2x = 62). These species hybridized and produced the amphidiploid Spartina X townsendii (2n = 2x = 62) which was sterile (Marchant 1963, 1966). But a fertile allopolyploid was also produced on the tidal mud flats at Southampton Water in the UK and this one, called S. anglica (2n = 4x = 120, 122, or 124), is fertile and can coexist with the sterile diploid. Isozyme evidence showed that S. anglica had very low genetic diversity, thus indicating it derived from a single hybridization event, or from multiple origins but from genetically uniform parents. Another hybrid, S. X neyrautii was discovered in 1892 in France. It has a similar isozyme profile as S. X townsendii but may represent the reciprocal hybrid. More recent work using DNA markers by Ayres and Strong (2001) confirms the origin of S. townsendii and S. anglica from the two purported parent species.

9. Tragopogon (goatsbeard, Asteraceae): Three diploid (all 2n = 2x = 12) European weedy species have been introduced to the U.S.: T. dubius, T. pratensis, and T. porrifolius. These species can each hybridize with the other, but the resulting F1s are sterile. Some fertile tetraploids are known (2N = 4X = 24). The fertile allotetraploids are morphologically distinct and reproductively isolated from the parents and have therefore been named T. mirus (T. dubius X T. porrifolius) and T. miscellus (T. dubius X T. pratensis) (Ownby, 1950).

10. Appalachian Spleenworts (Asplenium): Reticulate pattern of evolution involving hybridization and polyploidy well known and common in ferns. Asplenium species include diploids such as A. montanum, A. rhizophyllum, and A. platyneuron. These hybridize to form a complex of species, some of which are fertile allotetraploids (e.g. A. pinnatifidum) and others that are sterile triploids or apogamous hybrids.

11. Poa annua (2n = 4x = 28): Derived from a possible hybridization between the ephemeral species P. infirma (= P. exilis) of the Mediterranean and the perennial P. supina of mountainous areas. The two came into contact during the Pleistocene (Tutin, 1957). But - the karyotype does not have the pattern expected for the tetraploid derived from these two species. Maybe another Poa species is involved in producing P. annua.

12. Polypodium ferns: In Europe there are many variants of Polypodium vulgare. The tetraploid P. vulgare (AABB, 2N = 4X = 148) occurs and sometimes its range overlaps with the diploid P. australe (= P. cambricum, CC, 2N = 74). These hybridize and form a triploid (ABC). Also known is the allohexaploid P. interjectum (AABBCC, 2N = 6X = 222). Manton (1950) and Shivas (1961) looked at these ferns cytologically and synthesized the 5X pentaploid using the tetraploid P. vulgare and the hexaploid P. interjectum. This complex is tied to the North American Polypodium via hybridization and polyploidy. North American species include P. virginianum (4x), P. appalachianum (2x), and P. sibiricum (2x). The Flora North America treatment of Polypodium by Chris Haufler and colleagues (1993) shows the "reticulogram" that diagramatically shows relationships among the diploids, triploids and tetraploids.

Application of Allopolyploidy in Crop Improvement: 1. Bridging cross: Amphidiploids can be used as a bridge where direct cross between two species is not possible due to sterility in F1. Through this process the character from one species to a related species, generally from wild species to cultivated species. In such cases, at first an amphidiploid is made by chromosome doubling of an interspecific F1 hybrid, and then the amphidiploid is crossed with the recipient species. E.g., cotton, tobacco. In tobacco, Nicotiana digluta (Allohexaploid) is used as bridge in the transfer of tobacco mosaic virus resistance gene from Nicotiana sylvestris to Nicotiana tabacum.In cotton, an amphidiploid is used as a bridge for the transfer of gene from Gossypium thurberi to Gossypium hirsutum.

2. Creation of new crop species: Alloploidy sometimes helps in creation of new species as Triticale is the best example which is an alloploid between Triticum aestivum and Secale cereale. Triticale varities are mainly cultivated in Polland, Germany and France.Raphanobrassica, the triploid (AAC), was also an promising creation of new variety, which was of no use as the desired traits was not been obtained from the cross.

3. Interspecific Gene Transfer: In case of unavailability the desirable characters within the species, it is transferred from the related species. It is done in two ways as i) by alien addition or ii) by alien substitution. Such kind of gene transfer is found in wheat , tobacco, cotton and oats. In cotton, lint strength is transferred from Gossypium thurberi to Gossyium hirsutum

4. Tracing the origin of crop species: Alloploidy study is used to identify the origin of natural alloploidy plants.

Limitation of Alloploidy:

1. It is not possible to predict the effect of alloploids. 2. There may have defects in the newly synthesized alloploids. 3. It requires considerable time, labor and resources for the synthesis of alloploids through extensive breeding.

4. There are onely a small proportion of alloploids are promoising. 5. Chances of developing new species are very low.

Experimental Studies of Polyploids

A.        Crosses between diploids (AA, BB, or CC) and a tetraploid (AABB) would yield three different kinds of triploids (AAB, ABB, and ABC). In theory, the number of bivalents and univalents in the triploids could be used to reveal which diploid parent was involved.

B.                 Accidental polyploidy: Primula floribunda and P. verticillata were growing in a greenhouse at Kew Botanic Gardens. A fertile allotetraploid plant was found growing under the benches that was assumed to be a hybrid between these, named Primula kewensis . Some doubt expressed by later workers that this really was a hybrid derivative of these two species.

C.                 Resynthesis of wild polyploids: Müntzing (1930) interested in the origin of the mint Galeopsis tetrahit. He crossed Galeopsis pubescence (female) with Galeopsis speciosa (male). All were 2n = 2x = 16. The F1 was crossed to produce an F2 and the plants were quite variable morphologically and cytologically. One 2n = 3x = 24 (triploid) was formed from a reduced and unreduced gamete. The triploid was backcrossed to G. pubescens

(female) producing a plant with 2n = 4x = 32. This plant was morphological similar to G. tetrahit and it crossed with wild members of that species and produced fertile offspring. D. Bennett et al. (1992) used GISH to look at two grass species in the genus Milium to determine whether the diploid M. vernale (2n = 8) was one parent of the tetraploid M. montianum (2n=22). Wanted to know if the four large chromosomes seen in the M. montianum were derived from M. vernale. Extracted M. vernale genomic DNA, labeled it with biotin. Used this DNA as a probe on chromosome squashes of M. montianum. The biotinylated DNA was detected with fluorochrome-conjugated avidin whose signal was "amplified" with biotinylated antiavidin-D antibody. Chromosomes with positive hybridizations show up as yellow under U.V. light. Nonhybridizing portions of chromosomes counterstained red with propidium iodide. Yellow showed up only on the 4 large chromosomes, supporting concept they were derived from M. vernale. But note that some parts of the large chromosomes do not light up. These "repatterned" parts may have arisen after the formation of the allopolyploid or from limited transfer.

Conclusion:

 Polyploidy is a prominent force of shaping the evolution of plants (Winge 1917; Karpechenko, 1927; Stebbins, 1950, 1971), mostly in ferns (Wagner and Wagner, 1980; Werth et al., 1985, Vogel et al., 1999), and flowering plants (Soltis and Soltis, 2000; Wendel, 2000). Many of the crop plants are obviously polyploidy i.e., cotton, wheat, potato, alfalfa, etc. while the some others retain the vestiges of ancient polyploidy events (paleopolyploids) i.e., maize, soybean, cabbage etc. Though polyploidy got attraction initially due to their unique cytogenetics and their reproductive isolation(Blakeslee 1921, Jørgensen 1928), it was soon recognized that polyploids also exhibited distinctive phenotypic traits and hybrid vigor useful for agriculture (M¨untzing 1936, Randolph, 1941; Levin, 2002; Ramsey and Schemske, 2002). The polyploidy species formed independently from heterozygous diploid progenitors may provide important source of genetic variation (Soltis and soltis, 2000). Polyploids generally differ markedly from their progenitors in morphological, ecological,physiological and cytological characteristics (Levin 1983, 2002; Lumaret 1988) that can contribute both to exploitation of a new niche and to reproductive isolation.Thus, polyploidy is a major mechanism of adaptation and speciation in plants (Clausen et al., 1945; Stebbins, 1950; Grant, 1981; Otto & Whitton 2000; Levin, 2002). Already many of successful evidence have been developed. But there are still a lot of mystery are in the hide. It is the polyploidy breeding through which new crops can be developed and interspecific genes can be transferred and also the origin of crops can be traced. Though there is a lot of chances of revealing of undesirable characters and may have to face with several challenges, polyploidy breeding will reveal many of mysteries of the plant. It is now an interesting field of study to reveal the evolution of crop plants and utilizing their variability in the field of crop breeding.

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