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1

Obtaining carrot (Daucus carota L.) plants in isolated microspore cultures

100%
Gorecka K., Kowalska U., Krzyzanowska D., Kiszczak W.
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vol. 51
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issue 2
141-147
EN
Microspores were cultured on the modified B5 liquid medium containing 2.4D (0.1 mg L?1), NAA (0.1 mg L?1), L-glutamine (500 mg L?1), L-serine (100 mg L?1), and sucrose (100 g L?1). The developmental stages of microspores and divisions were observed. Initially, the formation of binuclear and multicellular structures was noticed. Plants regenerated in the cultures in which the tetrad stage of microsporogenesis had predominated. Embryoids were still forming 24 weeks after the cultures were set up. Six weeks after the transfer of androgenetic embryos onto the B5 regeneration medium, they were converted into complete plants. Out of 90 androgenetic plants planted in a growth chamber, 42 plants adapted to the new conditions. All of those plants proved to be diploids in cytometric analysis.
2

Instability of ploidy level during regeneration of transformed and non-transformed tobacco (Nicotiana tabacum L.)

86%
Sliwinska E., Zimny J., Drozdowska J.
Biotechnologia
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2003
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issue 3
260-266
EN
In plant tissue cultures, somaclonal variation is often observed. It can be an effect of the changes in the individual chromosome number or in the ploidy level. Flow cytometry, a fast and accurate method for the estimation of the nuclear DNA content, can be applied to study these changes. The DNA content in differentiated tissues of Nicotiana tabacum cultured in vitro was estimated using Partec CCA flow cytometer, starting from explant, through callus, up to regenerated shoots. The explant constituted stem segments of N. tabacum plants, non-transformed and transformed with gfp gene. Flow cytometric analysis showed differences in the proportion of 2C, 4C, 8C and 16C cells in plant tissue in different culture stages. Among the regenerated plantlets originated from non-transformed and transformed plants, diploid, tetraploid and mixoploid forms were observed. The transformation did not influence the share of cells representing different ploidy levels in the investigated plant material.
3

Application of flow cytometry for the study of plant genomes

86%
Dolezel J.
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vol. 38
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issue 3
285-302
EN
During the last fifteen years, flow cytometry and sorting greatly contributed to the improvement of our knowledge of plant genome structure and function. This paper reviews the applications of flow cytometry for the analysis of isolated nuclei and chromosomes. Because of its speed, precision and convenience, this method of analysis of the nuclear DNA content finds an enormous number of applications which cover basic research, breeding and production. The results obtained with chromosome analysis and sorting indicate that the technique might greatly simplify the analysis of plant genomes at the molecular level.
4

Karyological characterization of sugar beet gynogenetic lines cultured in vitro

86%
Svirshchevskaya A., Dolezel J.
Journal of Applied Genetics
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2001
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vol. 42
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issue 1
21-32
EN
Flow cytometry was used to screen ploidy levels in 47 cultured in vitro sugar beet gynogenetic lines of various origin and age, obtained after plant regeneration from unfertilized ovules. When donor plants were diploid, the majority of regenerants were found to have cells with 1C, 2C and 4C relative DNA content (mainly haploid and diploid) and there were large differences in the rate of spontaneous in vitro chromosome doubling between individual homozygous lines. Six ovule-derived lines regenerated from fertile and sterile diploid donors of forty-five lines were solid diploids from the very early stages of their in vitro cultivation, and thus could not be classified as doubled haploids. In the case of tetraploid donor plants, the gynogenetic regenerants demonstrated 2x-ploidy level. The results obtained in chimeric plants with both haploid and diploid cells indicated the possibility to overcome mixoploidy by their re-cultivation through generative shoot tip culture. The flow cytometry method confirmed data obtained by conventional microscopic chromosome counting in dividing leaf cells and was found very useful for screening of a large number of regenerants and for characterizing the process of in vitro gynogenetic lines formation in sugar beet.
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