**6. Monotelodisomics, monoisodisomics and haploids in cotton** *G. hirsutum* **L.**

Other type of chromatin deficiency namely monotelodisimics are characterized with absence of a chromosome arms. As a result 25 normal bivalents and one heteromorphic bivalent with arm deficiency formed in meiosis. There are 11 monotelodisomics following pollen and seed irradiation and 9 from different monosomic progenies in our collection at present time. Misdivision of a chromosome via centromere region followed by irradiation produced telocentric chromosome formation. Centromere inactivation caused loss of a chromosome arm. Telosome pair was detected as two different size univalents with various frequencies in the monotelodisomics (to 1.85±0.15 in average per cell). A high frequency of heteromorphic bivalents (to 0.95±0.05 in average per cell) was registered in monotelodisomics (Figure 22). Among the plants with arm deficiencies three had also a translocation.

**Figure 22.** Meiotic configurations in monotelodisomic plant in cotton *G. hirsutum*. Meiotic metaphase I cells showing 25 normal bivalents and 1 heteromorphic bivalent with arm deficiency in plant M1-1608/2. The arrow point out to the heteromorphic bivalent. Note that the background of figures was cleaned using Adobe Photoshop CSS extended ver‐ sion 12.

Monotelodisomics with the translocation had a telocentric chromosome as univalent in the majority of PMCs. As it is known, telocentrics for short arms are less often paired with normal homologous chromosomes [55]. Meiotic index was high (from 81.86±0.85 to 99.56±0.16), but the pollen fertility reduced in the monotelodisomics.

Isochromosome formation was also connected with damaging action of radiation to centro‐ mere chromosome regions. As a result of centromere inactivation instable telocentric formed its arm developed on 1800 , gave an isochromosome. Three monoisodisomics our collection were differed with isochromosome pairing at metaphase I of meiosis. If one of them had heteromorphic in most PMS (to 0.96±0.04) in other plants the isochromosome was often as univalent (to 0.91±0.22 an average on PMS). In spite of high Mi (to 98.55±0.25) in two monoi‐ sodicomics pollen fertility was reduced.

**Figure 21.** "Critical configurations" of the chromosomes at the meiotic metaphase I cells showing 23 bivalents and 1 univalent and one hexavalent (A) and 23 bivalents and 1 univalent and one quadrivalent (B-E) in cotton F1 plants from crosses the monosomics x translocation lines: (A) Mo13 xTr2; (B) Mo56 x Tr6, (C) Mo75 xTr16; (D) Mo77 x Tr21; (E) Mo77 x Tr25. The arrows point to the univalents, quadrivalents and hexavalent. Note that the background of figures

Other type of chromatin deficiency namely monotelodisimics are characterized with absence of a chromosome arms. As a result 25 normal bivalents and one heteromorphic bivalent with arm deficiency formed in meiosis. There are 11 monotelodisomics following pollen and seed irradiation and 9 from different monosomic progenies in our collection at present time. Misdivision of a chromosome via centromere region followed by irradiation produced telocentric chromosome formation. Centromere inactivation caused loss of a chromosome arm. Telosome pair was detected as two different size univalents with various frequencies in the monotelodisomics (to 1.85±0.15 in average per cell). A high frequency of heteromorphic bivalents (to 0.95±0.05 in average per cell) was registered in monotelodisomics (Figure 22).

**6. Monotelodisomics, monoisodisomics and haploids in cotton** *G.*

Among the plants with arm deficiencies three had also a translocation.

was cleaned using Adobe Photoshop CSS extended version 12.

*hirsutum* **L.**

278 World Cotton Germplasm Resources

Deficiencies for one chromosome arm occurred in the progenies of 9 monosomics. Thus, in four monosomic progenies (Mo2, Mo19, Mo34 and Mo61) that differed with respect to monosome transmission rates, monotelodisomics were produced due to univalent instability and resulted in misdivision. In the progenies of Mo6, Mo21, Mo22, Mo49, Mo54 and Mo68 daugher monosomics failed to produce, but monotelodisimics (from the progenies of Mo6, Mo21, Mo22, Mo49 and Mo68) and a monoisodisomic plant (from the progeny of Mo54) were detected. The results suggested an irregular univalent chromosome centromere misdivision in the parental monosomics that led to a single chromosome arm missing and formed either telocentric or isochromosome in the case of an arm doubling. Our results demonstrated the rather rare occurrence of telo-and isochromosomes in the monosomic progenies studied, which showed univalent misdivision to be rare.

Haploid plants of cotton are characterized by presence of 26 univalent chromosomes and by significant decrease in vigour and fertility, size of lives, bolls and flowers. Our collection includes four haploid plants. One haploid was obtained by irradiation of the seeds by fast neutrons (1005/22), other two (1579/6 and 171/417) – by pollen irradiation in M1 and M2 generations and one – from monosomic progeny (175/418-57). Meiosis was studied in the microsporocytes of the haploid plants 1005/22. Among 44 studied PMCs at metaphase I only five cells had open bivalents (to 0.14±0.06 on average per cells) (Figure 23 A). Others PMCs were characterized by presence of 26 univalents (Figure 23 B, C). As results, the range of polyads observed in PMCs.

**Figure 23.** Meiotic configurations in haploid plant 171/417 in cotton *G. hirsutum*. Meiotic metaphase I cells showing (A) 25 univalent and 1 bivalent; (B-C) 26 univalent. The arrow point out to the bivalent. Note that the background of figures was cleaned using Adobe Photoshop CSS extended version 12.

**Figure 24.** Meiosis in haploid plant 171/417 in cotton *G. hirsutum*. Abnormal sporads: (A) diad with micronuclei; (B)

Different mechanisms are proposed to explain the emergence of haploids during pollen irradiation. One of them suggests that haploids result from female parthenogenesis induced by pseudofertilization with irradiated pollen. According to another mechanism, fertilization occurs before zygotization, but the damaged paternal genome is eliminated early in develop‐ ment [56]. Some authors believe that haploids of *G. hirsutum* are completely sterile, wheareas haploids *G. barbadense* are fertile and produce seeds after pollination with normal pollen [57]. Moreover, a line of the latter species is known that frequently produced haploids of the

pentad with two micronucleis; (C) heksad with two micronucleis; (D and E)-oktads; (F) abnormal poliad.

**7. Storage and propagation cytogenetical collection of cotton**

Seeds Cytogenetical Collection of cotton maintained under room conditions (20-250

is no facility available for cold storage of seeds. They are placed in to parchment paper bags. Each bag has catalogue number and year of collection. Bags are stored in special metal boxes (30 x 11 cm) and boxes are placed in wooden-cases. Monosomic and translocation plants and

C). There

Cytogenetic Collection of Uzbekistan http://dx.doi.org/10.5772/58589 281

androgenous and matroclinous types.

All the 26 chromosomes remained as univalents in other two haploid plants (1579/6 and 175/418-57). Most abnormal haploid plant (1579/6) had low habit and a thin stem, compact bush, and scarce foliage. It was completely sterile. Its prominent feature was the complete absence of chromosome pairing. In all PMCs, meiosis studies revealed 26 univalents, scattered throughout the cells. Analysis of sporads revealed a significant decrease in meiotic index (to 18.97±1.31%), an increase in the number of tetrads with micronuclei (to 12.01±1.09%), and formation of abundant monads, dyads, triads, and polyads. The complete absence of chromo‐ some pairing resulted in the formation of imbalanced and abortive gametes and pollen sterility. Haploid plant 171/417 had rare open bivalents and one trivalent in several PMCs. Some PMSs of the two haploid had 26 bivalents. Non-disjunction during premeiotic mitosis could give rise to such a diploid cell.

Analysis of sporads revealed a significant decrease in meiotic index (to 13.12±0.92%), the decrease in the number of tetrads with micronuclei (to 3.02±0.46%), and formation of abundant pentads (16.51±0.01%), hexads (22.33±1.13%), heptads (17.17±1.02%) and oktads (10.46±0.83%) (Figure 24). In three years few bolls were obtained on this haploid plant (Figure 25).

five cells had open bivalents (to 0.14±0.06 on average per cells) (Figure 23 A). Others PMCs were characterized by presence of 26 univalents (Figure 23 B, C). As results, the range of

**Figure 23.** Meiotic configurations in haploid plant 171/417 in cotton *G. hirsutum*. Meiotic metaphase I cells showing (A) 25 univalent and 1 bivalent; (B-C) 26 univalent. The arrow point out to the bivalent. Note that the background of

All the 26 chromosomes remained as univalents in other two haploid plants (1579/6 and 175/418-57). Most abnormal haploid plant (1579/6) had low habit and a thin stem, compact bush, and scarce foliage. It was completely sterile. Its prominent feature was the complete absence of chromosome pairing. In all PMCs, meiosis studies revealed 26 univalents, scattered throughout the cells. Analysis of sporads revealed a significant decrease in meiotic index (to 18.97±1.31%), an increase in the number of tetrads with micronuclei (to 12.01±1.09%), and formation of abundant monads, dyads, triads, and polyads. The complete absence of chromo‐ some pairing resulted in the formation of imbalanced and abortive gametes and pollen sterility. Haploid plant 171/417 had rare open bivalents and one trivalent in several PMCs. Some PMSs of the two haploid had 26 bivalents. Non-disjunction during premeiotic mitosis could give rise

Analysis of sporads revealed a significant decrease in meiotic index (to 13.12±0.92%), the decrease in the number of tetrads with micronuclei (to 3.02±0.46%), and formation of abundant pentads (16.51±0.01%), hexads (22.33±1.13%), heptads (17.17±1.02%) and oktads (10.46±0.83%)

(Figure 24). In three years few bolls were obtained on this haploid plant (Figure 25).

figures was cleaned using Adobe Photoshop CSS extended version 12.

polyads observed in PMCs.

280 World Cotton Germplasm Resources

to such a diploid cell.

**Figure 24.** Meiosis in haploid plant 171/417 in cotton *G. hirsutum*. Abnormal sporads: (A) diad with micronuclei; (B) pentad with two micronucleis; (C) heksad with two micronucleis; (D and E)-oktads; (F) abnormal poliad.

Different mechanisms are proposed to explain the emergence of haploids during pollen irradiation. One of them suggests that haploids result from female parthenogenesis induced by pseudofertilization with irradiated pollen. According to another mechanism, fertilization occurs before zygotization, but the damaged paternal genome is eliminated early in develop‐ ment [56]. Some authors believe that haploids of *G. hirsutum* are completely sterile, wheareas haploids *G. barbadense* are fertile and produce seeds after pollination with normal pollen [57]. Moreover, a line of the latter species is known that frequently produced haploids of the androgenous and matroclinous types.

## **7. Storage and propagation cytogenetical collection of cotton**

Seeds Cytogenetical Collection of cotton maintained under room conditions (20-250 C). There is no facility available for cold storage of seeds. They are placed in to parchment paper bags. Each bag has catalogue number and year of collection. Bags are stored in special metal boxes (30 x 11 cm) and boxes are placed in wooden-cases. Monosomic and translocation plants and

of their hybrids are grown at the greenhouse conditions in soil. All data collected are stored as a hard copy catalogue book that is being conversed to electronic format.

**9. Conclusions**

germplasm exchange program.

**Acknowledgements**

in design of the chapter.

Laboratory, Uzbekistan

**Author details**

In conclusion we studied new Cotton Cytogenetic Collection adapted to the Central Asian condition in contrast Cytogenetic Collection from USA using different types of seed and pollen irradiation. We propose the presence of unique cotton aberrations involved chromosomes for absent chromosomes in American collection. The results suggested a detection of "reduced" stigma as a useful phenotypic marker for cotton monosomics which makes it possible to distinguish different cytotypes without cytological analyses. The results demonstrated of new unique desynaptic cotton plants in which progeny produced monosomics with high frequen‐ cy. We observed the very occurrence of univalents misdivision probably owing to monosome stability in the unique genetic background. Our cotton monosomic lines are unique and should be a valuable cytogenetic tool not only for chromosome assignment of new marker genes and genome enrichment with new chromosome deficient plant, but also for a development of new

Cytogenetic Collection of Uzbekistan http://dx.doi.org/10.5772/58589 283

Alternatively, the creation of chromosome substitution lines through crossing of each of the new monosomics with *G.barbadense* genotype (Pima 3-79) is in progress. This will serve as a foundation to apply molecular marker (e g., SSPs) for the identification of our monosomics in hybrids with chromosome substitutions for a given monosome. At the same time, our monosomic cotton lines with initial cytogenetic characteristics, which developed using single genome background, should be useful germplasm for cotton researchers to use as material for future breeding genetic, cytogenetic and molecular-genetic investigation of cotton genome. In future we plan to identify the chromosome deficiencies by molecular markers (SSR) to map of cotton genome. Also we will continue identification monosomic lines of our cytogenetic collection using a well-defined tester-set of translocation lines of the USA Cytogenetic Collection, kindly provided by Dr. D.M. Stelly, Texas A&M University, USA, under USDA

This work was partially supported by research grants 38/96, 28/98, 26/2000, F.4.15 and F-5-31 from Committee for Coordination of Science and Technology Development (CCSTD) of the Republik of Uzbekistah. We thank Dr. Svetlana Polyarush and Artyom Maknyov for their help

Marina Sanamyan, Julia Petlyakova, Emma Rakhmatullina and Elnora Sharipova

National University of Uzbekistan, Department of Biology and Soil, Cotton Genetics

cotton chromosome substitution lines and germplasm introgression.

## **8. Location, maintenance and funding**

The Cytogenetical Collection of cotton currently stored in the National University of Uzbe‐ kictan at Tashkent. It is funded by Committee for Coordination of Science and Technology Development (CCSTD) under the Cabinet Ministry of Uzbekistan.

**Figure 25.** Haploid plant 171/417 in cotton *G. hirsutum* obtained in M2 after pollen irradiation. (A) haploid plant; (B) fertile branch with two bolls; (C) leaf, flower and bract in parental line L-458 (1-3) and haploid plant (4-6); (D) cup, petal and staminate column in parental line L-458 (1-3) and haploid plant (4-7).
