BIOTECHNOLOGICAL METHODS - A TOOL FOR CREATING NEW FLAX GENOTYPES RESISTANT TO ANTHRACNOSE

2020 ◽  
Keyword(s):  
Culture Filtrate ◽  
Embryo Culture ◽  
Selective Medium ◽  
Field Resistance ◽  
Immature Embryos ◽  
Cell Selection ◽  
Morphogenic Callus ◽  
Morphogenetic Potential ◽  
Regenerant Plants

The purpose of this research was to create new flax genotypes resistant to anthracnose using biotechnological techniques and methods. As a result of studies with using a culture of immature embryos and a selective medium, flax regenerant plants resistant to the culture filtrate (CF) of the fungus - anthracnose pathogen Colletotrichum lini Manns et Bolley and line 21 resistant to this pathogen were obtained. A scheme for differentiating flax genotypes in vitro by resistance to anthracnose has been developed. It was established that upon cultivation of immature embryos on Sh-2 medium containing CF of the fungus, the causative agent of anthracnose, of a mixture of strains 680, 677 * 674, 674 * at a concentration of 36.0 ml / l, the amount of morphogenic callus formed in the first and second passages, expressed in percent, and the indicator of the field resistance of this genotype to anthracnosis on an artificial infectious-provocative background was close in value, and the number of morphogenic callus formed in the first and second passages can be used to judge the resistance of the studied genotypes to ant cancer and differentiate them by resistance to this pathogen. The influence of the flax genotype on the potency of cells to morphogenesis under selective conditions was revealed. Cells of genotypes L 957-8-7, Alexim, Punjab, Zaryanka had high morphogenetic activity. The morphogenetic potential of genotypes L 1506-8-4, Rosinka has already been exhausted by 2 ... 3 passages. It was found that biotechnological methods: cell selection in vitro, embryo culture are effective in creating genotypes of flax, more resistant to anthracnose than original forms.

scholarly journals OPTIMIZATION OF IN VITRO SELECTIVE MEDIA FOR SELECTING ANTHRACNOSIS-RESISTANT FLAX CELLS

2021 ◽  
Author(s):  
Н.В. Пролётова ◽  
Л.П. Кудрявцева
Keyword(s):  
Culture Filtrate ◽  
In Vitro Selection ◽  
Selective Medium ◽  
Selective Media ◽  
Primary Roots ◽  
Morphogenic Callus ◽  
Weakly Virulent ◽  
Germinated Seeds ◽  
Selection Of

Цель исследований – оптимизация селективных сред для проведения отбора in vitro каллусных клеток льна, устойчивых к культуральному фильтрату штаммов возбудителя антракноза и создание in vitro новых генотипов, устойчивых к болезни. В результате исследований уточнен состав культурального фильтрата штаммов антракноза. Выявлено, что токсичность культуральных фильтратов не зависела от вирулентности используемых штаммов – более токсичными оказались культуральные фильтраты штаммов 784 (сильновирулентного) и 780 (средневирулентного) (загнивание и отмирание первичных корешков на 5 сутки наблюдали у 67 – 88% проросших семян), менее токсичны – штаммы 793 (сильновирулентный) и 788 (слабовирулентный) (на 5 сутки загнивание и отмирание первичных корешков отмечено у 9 – 15% проросших семян). Установлено, что морфогенные очаги формировались активнее у генотипов, морфогенный каллус которых переносили на среду с аналогичной или более высокой концентрацией культурального фильтрата. Показано, что на 14 сутки во втором пассаже с большей частотой формировались морфогенные каллусы, почки и побеги при использовании в первом и втором пассажах селективной среды, содержащей культуральный фильтрат в концентрации 40 мл/л, или в первом пассаже – 40 мл/л, а во втором – 44 мл/л. Выделены генотипы, сохраняющие устойчивость к антракнозу в течение трёх поколений на уровне 50 – 60%: НО-78 х Ленок, HJI-103-2 х Ленок, НЛ-40-1 х Ленок, HЭ-38 х Росинка, НЭ-36 х Ленок, НЭ-17 х Ленок, HЭ-16-2 х Росинка. Research objective – optimization of selective media for in vitro selection of flax callus cells resistant to culture filtrate of anthracnose pathogen strains and in vitro creation of new disease-resistant genotypes. As a result of the research, the composition of the culture filtrate of anthracnose strains was clarified. It was revealed that the toxicity of cultural filtrates did not depend on the virulence of the strains used - cultural filtrates of strains 784 (highly virulent) and 780 (medium virulent) turned out to be more toxic (decay and death of primary roots on day 5 was observed in 67 - 88% of germinated seeds), less toxic - strains 793 (strongly virulent) and 788 (weakly virulent) (on the 5th day, decay and death of primary roots was noted in 9-15% of germinated seeds). It was found that morphogenic foci were formed more actively in genotypes, the morphogenic callus of which was transferred to a medium with a similar or higher concentration of the culture filtrate. It was shown that on the 14th day in the second passage, morphogenic callus, buds and shoots were formed with a greater frequency when using in the first and second passages a selective medium containing a culture filtrate at a concentration of 40 ml/l, or in the first passage - 40 ml/l, and in the second - 44 ml/l. Genotypes were identified that retain resistance to anthracnose for three generations at a level of 50 - 60%: NO-78 x Lenok, HJI-103-2 x Lenok, NL-40-1 x Lenok, NE-38 x Rosinka, NE-36 x Lenok, NE-17 x Lenok, NE-16-2 x Rosinka.

scholarly journals Improvement of the technology of obtaining stable (di)haploid regenerants from embryonic culture of apomictic sugar beet (Beta vulgaris) breeding material without the use of colchicine

2019 ◽  
Vol 6 (2) ◽  
pp. 3-17
Author(s):  
N. Kovalchuk ◽  
M. Roik ◽  
Ya. Hadzalo ◽  
T. Nediak ◽  
O. Zinchenko
Keyword(s):  
Sugar Beet ◽  
Seed Production ◽  
Embryo Culture ◽  
Direct Regeneration ◽  
Genetic Determination ◽  
Alloplasmic Lines ◽  
Cultural Medium ◽  
Wide Range ◽  
Regenerant Plants

Aim. To evaluate the effi ciency of inducing generative, reduced parthenogenesis and to better use the differentiating potential of the embryo culture under apomictic seed production in selection materials of sugar beet with cytoplasmic male sterility (CMS), and B) to isolate homozygous lines (dihaploids) without the use of polyploidizing substances. Methods. Apomictic (agamosper- mous) seed production in apocarpous pollen sterile lines from B. vulgaris subsp. vulgaris var. altissima (sugar beet) using classi- cal so-called Owen sterile cytoplasm and sterile cytoplasm from Beta maritimа and Beta patula as sources, was conducted under pollen free conditions and spatial isolation in the greenhouse breeding complex of the Yaltushkivska experimental breeding station (Yaltushki, Ukraine). The specifi cities of embryonic development of apomictic embryos were studied with the purpose of effi cient regulation of the induction of explants in vitro as donors of the culture of immature embryos. Fluorescent fl ow cytophotometry in combination with the computer program of the Partec Ploidy Analyser PA-2 (Partec GmbH, Germany, now Sysmex), were used to determine the degree of ploidy, enabling the selection of haploid and dihaploid lines in vitro. A genetic method was developed using the expression of morphological marker indices of nuclear genes of anthocyanin coloring (R+ r–) of regenerant plants in vitro and ploidy determination for differentiation by generative (reduced) parthenogenesis. The sampling technique that took into account the hormonal composition of cultural media and the level of genome ploidy, sample frequency and statistical analysis of the results was determined using the appropriate statistics; the percentage of regenerants, induced by different types of morpho- genesis and ploidy in vitro, was determined along with the measurement error to control the accuracy of the selected sampling (number of seed embryos). Results. The selected cultural medium No. 3, based on the basal medium according to Gamberg et al., 1968 (21), contained 6 BAP – 2 mg/l, 2.4 D – 0.5 mg/l, gibberellic acid – 0.1 mg/l, which ensured a success rate of 4.4 to 23.3 % of direct regeneration of shoots from the embryo culture, depending on the genotype of donors, and 4–10 % for induction and proliferation of callus. In ten experimental numbers of alloplasmic lines of sugar beet, the incidence of haploids and mixoploids among the regenerants from the embryo culture fl uctuated within the wide range of 14.8 – 62.2 % and exceeded the indices, ob- tained by other known methods of haploid parthenogamy, which had the values of 3.79 – 6.25 %. Conclusions. The homozygous lines and dihaploids were determined and set apart/stabilized in the process of micropropagation, where the differentiation of clones was made on the basis of total DNA content in interphase nuclei, using information of histograms generated in fl uorescent fl ow cytometry with the Partec Ploidy Analyser PA-II instrumentation. The medium, based on macro- and microsalts according to Gamberg et al., 1968 (21) was found to be the most effi cient; it ensured at least partially successful direct regeneration in the culture of embryos within the range of 4.40 ± 1.29 to 23.3 ± 3.45 %. The success of direct regeneration of apomictic material depended on the composition of the cultural medium used fi rst and foremost, and to a lesser extent on the stages of embryogenesis from day 12 till day 32, differentiated by the fi xation period for seed embryos starting from the beginning of fl owering. Homozygous lines were created without polyploid-inducing substances due to spontaneous transfer of some cells of haploid regenerant plants to a higher level of ploidy, that can be used in the breeding of sugar beet. Genetic determination of apomictic seed reproduction in alloplasmic lines and pollen free lines of sugar beet and the technologies of inducing dihaploids allow reducing the period of inzucht-crossing considerably to obtain homozygous lines, creating unique material for chromosome engineering and marker-oriented selection with target combinations of genes in homozygous state.

Paucity of somaclonal variation from immature embryo culture of barley

1989 ◽  
Vol 40 (6) ◽  
pp. 1155 ◽  
Author(s):  
DJ Luckett ◽  
D Rose ◽  
E Knights
Keyword(s):  
Tissue Culture ◽  
Somaclonal Variation ◽  
Embryo Culture ◽  
Field Trial ◽  
Immature Embryo ◽  
Immature Embryos ◽  
Golden Promise ◽  
Immature Embryo Culture ◽  
Tissue Origin ◽  
Regenerant Plants

Intact immature embryos of barley (cv. Golden Promise) and component tissues (the scutellum and embryonic axis) were cultured to produce callus. Regenerant plants were obtained from this callus and SC2 families raised. These families were examined in a field trial to search for somaclonal variation. No obvious variants were found confirming our previous unpublished results. The lack of somaclonal variation generated by barley tissue culture (which is in contrast to other species) was not a result of the tissue origin of the regeneration event.

scholarly journals USAGE OF CULTURAL FILTRATES IN IN VITRO SELECTION FOR ANTHRACNOSE RESISTANCE

2021 ◽  
Vol 212 ◽  
pp. 92-97
Author(s):  
N. V. Proletova ◽  
Keyword(s):  
In Vitro Selection ◽  
Selective Medium ◽  
Federal State ◽  
Scientific Center ◽  
Morphogenic Callus ◽  
Weakly Virulent ◽  
Vitro Development ◽  
Highly Virulent ◽  
Germinated Seeds

The research was carried out on the basis of the laboratory of selection technologies of the Federal State Budgetary Scientific Institution “Federal Scientific Center of Fiber Crops” (Tver region) in 2018–2020. The aim of the research is in vitro development of new flax genotypes resistant to anthracnose, one of the most harmful fungal diseases. As a result of the research, the composition of the cultural filtrate of the anthracnose causative agent was clarified. It was revealed that toxicity of the cultural filtrates did not depend on the virulence of the strains used in the present studies, the cultural filtrates of strains 784 (highly virulent) and 780 (medium virulent) turned out to be more toxic (decay and death of radicle was observed on the 5th day in 67 - 88% of germinated seeds), less toxic are strains 793 (highly virulent) and 788 (weakly virulent) (decay and death of radicle was observed on the 5th day in 9-15% of germinated seeds). It was found that morphogenic foci were formed more actively in genotypes the morphogenic callus of which was transferred to a medium with a higher concentration of the cultural filtrate; it was shown that in the second passage, when transferring morphogenic calli from a selective medium, which contains 40 ml / L of cultural filtrate on a selective medium also containing 40 ml / L of cultural filtrate, as well as on a selective medium containing 44 ml / L of cultural filtrate, the number of formed morphogenic calli and green buds on the 14th day is significantly higher than in case of transferring on a selective medium containing 36 ml / L of cultural filtrate. Viable regenerant plants were obtained and genotypes were isolated, which retained resistance to anthracnose for three generations at a level of 50 - 60%: NO-78 x Lenok, NL-103-2 x Lenok, NL-40-1 x Lenok, NE-38 x Rosinka, NE-36 x Lenok, NE-17 x Lenok, NE-16-2 x Rosinka.

scholarly journals LABORATORY EVALUATION OF REGENERATES OF WHEAT HYBRID COMBINATIONS IN VITRO AND EX VITRO CONDITIONS

ÈKOBIOTEH ◽  
2021 ◽  
Vol 4 (2) ◽  
pp. 81-88
Author(s):  
A.E. Zinatullina ◽  
◽  
V.I. Nikonov ◽  
Keyword(s):  
Embryo Culture ◽  
Histological Analysis ◽  
Food Resource ◽  
Selective Medium ◽  
Stress Factors ◽  
High Yield ◽  
Laboratory Evaluation ◽  
Ex Vitro ◽  
Culture In Vitro

Drought is the combination of climatic conditions that leads to a long-term shortage of water in the soil and air. This is one of the most common abiotic stress factors that leads to significant losses of crop yield and the emergence of a threat to food security. Researchers are actively developing ways to create drought-resistant zoned varieties of economically important agricultural crops and especially cereals as the main food resource. Such varieties should maintain a relatively high yield rate with a shortage of water in the soil and air. The aim of the work was the laboratory evaluation in vitro and ex vitro of wheat regenerants formed in the embryo culture in vitro under conditions selective for the indicator "drought resistance". Methods of embryo culture in vitro, laboratory evaluation of caryopsis viability, histological analysis, as well as statistical processing of the received results were used. Under the conditions of in vitro experiments on the selective medium simulating drought by introducing mannit at the concentration of 8% as an osmotic, regenerants of 5 hybrid wheat combinations that showed tolerance to stress were obtained. It is shown that the development of regenerants in vitro and ex vitro pass according to the same phenological phases and in the same duration as donor plants. Regenerants form caryopsises of sufficiently high quality, which is confirmed by laboratory observations of their viability and histological analysis of seedlings.

In Vitro Cell Selection of Sugar Beet for Resistance to Culture Filtrate of the Fungus Fusarium oxysporum

2019 ◽  
Vol 53 (4) ◽  
pp. 307-314
Author(s):  
R. S. Yerzhebayeva ◽  
A. M. Abekova ◽  
G. H. Bersimbaeva ◽  
K. T. Konysbekov ◽  
S. O. Bastaubaeva ◽  
...  
Keyword(s):  
Sugar Beet ◽  
Fusarium Oxysporum ◽  
Culture Filtrate ◽  
Cell Selection ◽  
Selection Of

scholarly journals Experimental study of embryo differentiation in angiosperms

2014 ◽  
Vol 50 (1-2) ◽  
pp. 257-263 ◽  
Author(s):  
T. B. Batygina ◽  
V. E. Vasilyeva
Keyword(s):  
Experimental Study ◽  
Embryo Culture ◽  
Sharp Increase ◽  
Outer Integument ◽  
Dry Weight ◽  
Critical Stage ◽  
Self Sufficiency ◽  
Morphogenetic Potential

By comparison of peony and lotus embryos in <em>in vitro</em> culture we established differences in their morphogenetic potential. We traced in peony embryo culture: embryogenesis, embryoidogenesis, organogenesis and histogenesis, whereas the lotus embryo enhibited only one pathway of development - embryogenesis. At the critical stage the embryo becomes independent of the maternal organism, i.e, self-sufficient. Self-sufficiency of the lotus embryo on the 4O-11th day after pollination correlates with development and greening of the first -lead in the plumule, with a sharp increase in embryo dry weight and content of carbohydrates, decrease in the size of the endosperm, degeneration of the inner integument, appearance of starch in the embryo and ovary walls an Increase of its content in the outer integument.

scholarly journals USE OF SUCROSE AND MANNITOL FOR DIFFERENTIATION OF FLAX GENOTYPES BY RESISTANCE TO OSMOTIC STRESS

2020 ◽  
Vol 15 (3) ◽  
pp. 10-15
Author(s):  
Elena Vinogradova
Keyword(s):  
Osmotic Stress ◽  
Sucrose Solution ◽  
Primary Root ◽  
Callus Cultures ◽  
Immature Embryos ◽  
Selective Agent ◽  
Scientific Center ◽  
Morphogenic Callus ◽  
Callus Tissues

The studies were carried out with the aim of studying the effect of various concentrations of sucrose and mannitol on seeds, immature embryos, and callus cultures of flax to develop a method for obtaining genotypes resistant to osmotic stress. The work was carried out in the Tver region in the laboratory of breeding technologies in 2017–2019. Flax varieties Barbara, Belinka, LM-98, Aurore, Tverskoy, Svetoch, Diplomat, Symfonia were used as objects of research. The seeds were obtained from the National Flax Collection of the Federal Scientific Center for Bast Crops. The effect of sucrose solution on the length of the primary root was detected at concentrations - 0; 8.7; 14.9%. To assess the germination energy of seeds under osmotic stress, the concentration of sucrose was reduced and the range 0 (control) ... 9% was considered. Immature embryos removed from the capsules on the 10th day after pollination were cultivated on MS medium with sucrose, as a selective agent, at a concentration of 5.0 ... 7.0%. Callus tissues were cultured using mannitol as an osmotic at concentrations of 0; 30.0; 36.4; 37.0; 37.4; 38.0 mg/l. Concentrations of 5.0, 6.0 and 7.0% of sucrose can be used as an osmotic differentiator for seeds (10 ... 80% of seeds germinated in the Belinka variety, 80 ... 100% in the Varbara variety, 80 ... 90% in the variety LM-98). Sucrose, as a selective agent, in a culture of immature flax embryos in vitro at a concentration of 5.0 ... 7.0% can be selective only for certain genotypes, for example, the Aurore variety. The selection of resistant callus cells, followed by the formation of adventive buds and shoots in the meristematic foci, can be carried out on media containing 30.0 or 36.4 mg / L of osmosis, which allows obtaining morphogenic callus, buds, shoots in all studied genotypes, as well as in the Aurore variety 1.1 ... 1.2 byp./callus, in the Tverskoy variety - 0.6 ... 0.8, in the Barbara variety - 1.0 ...1.1

Cell selection of wheat in vitro for heat resistance and the effect of endophytic infection on morphogenetic potential.

Biomics ◽  
2018 ◽  
Vol 10 (1) ◽  
pp. 33-36
Author(s):  
M.H. Mamedova ◽  
F.A. Alisoy ◽  
C.M. Talai ◽  
E.R. Ibragimov ◽  
T.H. Karagozov
Keyword(s):  
Heat Resistance ◽  
Cell Selection ◽  
Morphogenetic Potential ◽  
Selection Of

scholarly journals In Vitro Culture of Immature Zygotic Mango Embryos and Plantlet Development

HortScience ◽  
2011 ◽  
Vol 46 (11) ◽  
pp. 1528-1532 ◽  
Author(s):  
Juan Bernardo Pérez-Hernández ◽  
María José Grajal-Martín
Keyword(s):  
In Vitro Culture ◽  
Embryo Culture ◽  
Culture System ◽  
Zygotic Embryo ◽  
Hybrid Plant ◽  
Coconut Water ◽  
Immature Embryos ◽  
Culture Techniques ◽  
Total Treatment

In vitro culture of immature embryos may assist mango breeding in the production of hybrid plant material. However, zygotic embryo culture techniques have not been successfully developed for mango. To recover in vitro zygotic plants through embryo culture, ‘Lippens’ and ‘Keitt’ were used as a source of model immature embryos. Excised embryos were incubated in a liquid maturation medium to test different culture systems and media composition. Subsequent germination allowed for the recovery of complete in vitro plantlets. Variables included during artificial embryo maturation, independently or through paired interactions, significantly affected all the parameters measured for embryo development and characterization of the plantlets. Main effects of culture system (i.e., static versus agitation) and coconut water supply (20%) were responsible for up to 85.5% of total treatment variation. Direct and inverse interactions observed between culture system and either coconut water supplement or sucrose content (45 or 60 g·L−1) contributed to define the best combination of factors to improve embryo growth and plant formation. Complete plantlets could be obtained at a frequency above 83% for both cultivars at the end of the in vitro phase at a developmental stage that allowed acclimatization to greenhouse conditions.

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