Молекулярно-генетичні механізми стійкості до холодового стресу в геномно-заміщеної форми пшениці: автореферат дисертації на здобуття наукового ступеня кандидата біологічних наук
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Date
2017
Authors
Єфіменко, Тетяна
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Автореферат дисертації на здобуття наукового ступеня кандидата біологічних наук за спеціальністю 03.00.15 – генетика. – Інститут клітинної біології та генетичної інженерії НАН України, Київ, 2017.
В дисертації показано зв’язок характеристик кореневої системи з підвищеної зимостійкістю для амфідиплоїда Авротіка (ААВВТТ) та інтрогресивних ліній пшениці з генетичним матеріалом Т геному виду Aegilops mutica. Створені інтрогресивні лінії, що походять від схрещування сорту м’якої пшениця Аврора і амфідиплоїда Авротіка, оцінено з використанням мікросателітного аналізу щодо присутності інтрогресій у складі хромосом 5 гомеологічної групи. Показано присутність "мозаїчних" хромосом, що могли утворитися через рекомбінації геномів D і Т. Визначено, що накопичення вільного проліна та зміни проникності мембрани не належать до механізмів забезпечення підвищеної зимостійкості амфідиплоїда Авротіка. Встановлено позитивний зв’язок між об’ємом кореневої системи Авротіки та інтрогресивних ліній та виживанням рослин у польових умовах зимою. Показано, що характерний для Авротіки алель гена AGL21, продукт якого регулює синтез ауксина в коренях, переважає у рослин, що перезимували в полі, отже, може мати позитивний зв’язок з морозостійкістю рослин.
A thesis abstract submitted to acquire the degree of Doctor of Philosophy (PhD) in Biology, speciality 03.00.15 – genetics. – Institute of Cell Biology and Genetic Engineering of National Academy of Sciences of Ukraine, Kyiv, 2017. The dissertation is devoted to the investigation of mechanisms of higher winter hardiness of the amphidiploid Aurotica, which combines in its genome AABB tetracomponent of common wheat cultivar Aurora and TT genome of the diploid wild wheat relative Aegilops mutica (Amblyopyrum muticum). Introgressive lines Triticum aestivum / Aegilops mutica were developed by crossing Aurora x Aurotica and the following self-pollination of the hybrids, which was possible due to the recombination between the chromosomes of D and T genomes. Chromosome numbers were determined in hybrid generations F3, F4 and F5, and only 40-43-chromosome F4 plants were chosen to form F5 generation in order to get 42-chromosome euploids introgressive lines. Changes in chromosome plates of the hybrid plants were observed: presence of telocentric chromosomes, dicentric chromosomes and number o satellite chromosomes differing from Aurora and Aurotica, these changes could be evidence of chromosome rearranges in the genomes of hybrid plants possibly caused by introgression of alien genetic material. Presence of T genome genetic material in the genomes of the introgressive lines was confirmed by dot blot hybridization with genome DNA of Aegilops mutica as probe; most lines demonstrated positive hybridization signals. Additionally, morphological traits assessment also gave evidence on the presence of alien genetic material in the genomes of the lines: all lines differed from Aurora at least for some traits that have been assessed (traits of the spike and vegetative part of the plant). In order to indentify introgressions in chromosomes of homeological group 5, which according to the literature contains main genes for freezing tolerance, microsatellite analysis was conducted with SSR loci specific for this group of chromosomes. According to the results of the SSR analysis, it could be assumed that some wheat chromosomes of homeological group 5 were replaced by the chromosomes of T genome; other chromosomes had replaced arms or smaller translocations, which could form “mosaic” chromosomes with alleles at some SSR loci specific to Aurora, and at other SSR loci – alleles specific to Aurotica. It was shown that lines with alleles of SSR loci on 5A chromosome specific to Aurotica were characterized by better winter field survival. According to the results of free proline quantification and determination of electric conductivity (electrolyte leakage), it was demonstrated that Aurotica accumulated less free proline during the period of cold acclimation and was characterized by the higher electric conductivity, compared to Aurora. These results indicated that higher winter hardiness of Aurotica was provided by different molecular mechanisms than free proline accumulation and changes in membrane that prevent electrolyte leakage after freezing stress. In the experiments of laboratory freezing of Aurora and Aurotica seedlings (crowns) and the following survival assessment, Aurotica demonstrated better survival. During the preparation for the freezing experiments, when seedlings were excavated from the soil after acclimation, a useful observation was made that Aurotica formed more developed (stronger) root system compared to Aurora when grown at low positive temperature (4°C). Based on this observation, the next root system characteristics were studied for Aurotica, Aurora and introgressive lines. For the root system study plants were grown in the paper boxes in laboratory and also in the field. Such root system characteristics as maximum root length, root number and root volume (determined as volume of liquid supplanted by the roots) were determined. Difference between Aurora and Aurotica was identified for root volume. It was shown that introgressive lines with higher correlation of root volume to the number of productive shoots were characterized by better field survival. Primers were developed to the coding sequence of the gene AGL21, which regulates root development via positive regulation of auxin accumulation in lateral root primordia. Aurora and Aurotica had polymorphic products with primers to second part of the gene. Introgressive lines were analyzed, and it was demonstrated that lines with AGL21 allele specific for Aurotica developed root system with greater volume at cold acclimation conditions, and were characterized by better winter field survival.
Автореферат диссертации на соискание ученой степени кандидата биологических наук по специальности 03.00.15 – генетика. – Институт клеточной биологии и генетической инженерии НАН Украины, Киев, 2017. В диссертации показано связь характеристик корневой системы с повышенной зимостойкостью для амфидиплоида Авротика (ААВВТТ) и интрогрессивных линий пшеницы с генетическим материалом генома Т вида Aegilops mutica. Созданные интрогрессивные линии, которые происходят от скрещивания сорта мягкой пшеницы Аврора и амфидиплоида Авротика, оценены с использованием микросателитного анализа относительно присутствия интрогрессий в составе хромосом 5 гомеологической группы. Показано присутствие "мозаических" хромосом, которые могли возникнуть в результате рекомбинации геномов D и Т. Определено, что накопление свободного пролина и изменения проницаемости мембраны не относятся к механизмам обеспечения высокой зимостойкости амфидиплоида Авротика. Установлено позитивную связь между объемом корневой системы Авротики и интрогрессивных линий и выживанием растений в полевых условиях зимой. Показано, что характерный для Авротики аллель гена AGL21, продукт которого регулирует синтез ауксина в корнях, встречается чаще у растений, которые перезимовали в поле, следовательно, может иметь связь с зимостойкостью растений.
A thesis abstract submitted to acquire the degree of Doctor of Philosophy (PhD) in Biology, speciality 03.00.15 – genetics. – Institute of Cell Biology and Genetic Engineering of National Academy of Sciences of Ukraine, Kyiv, 2017. The dissertation is devoted to the investigation of mechanisms of higher winter hardiness of the amphidiploid Aurotica, which combines in its genome AABB tetracomponent of common wheat cultivar Aurora and TT genome of the diploid wild wheat relative Aegilops mutica (Amblyopyrum muticum). Introgressive lines Triticum aestivum / Aegilops mutica were developed by crossing Aurora x Aurotica and the following self-pollination of the hybrids, which was possible due to the recombination between the chromosomes of D and T genomes. Chromosome numbers were determined in hybrid generations F3, F4 and F5, and only 40-43-chromosome F4 plants were chosen to form F5 generation in order to get 42-chromosome euploids introgressive lines. Changes in chromosome plates of the hybrid plants were observed: presence of telocentric chromosomes, dicentric chromosomes and number o satellite chromosomes differing from Aurora and Aurotica, these changes could be evidence of chromosome rearranges in the genomes of hybrid plants possibly caused by introgression of alien genetic material. Presence of T genome genetic material in the genomes of the introgressive lines was confirmed by dot blot hybridization with genome DNA of Aegilops mutica as probe; most lines demonstrated positive hybridization signals. Additionally, morphological traits assessment also gave evidence on the presence of alien genetic material in the genomes of the lines: all lines differed from Aurora at least for some traits that have been assessed (traits of the spike and vegetative part of the plant). In order to indentify introgressions in chromosomes of homeological group 5, which according to the literature contains main genes for freezing tolerance, microsatellite analysis was conducted with SSR loci specific for this group of chromosomes. According to the results of the SSR analysis, it could be assumed that some wheat chromosomes of homeological group 5 were replaced by the chromosomes of T genome; other chromosomes had replaced arms or smaller translocations, which could form “mosaic” chromosomes with alleles at some SSR loci specific to Aurora, and at other SSR loci – alleles specific to Aurotica. It was shown that lines with alleles of SSR loci on 5A chromosome specific to Aurotica were characterized by better winter field survival. According to the results of free proline quantification and determination of electric conductivity (electrolyte leakage), it was demonstrated that Aurotica accumulated less free proline during the period of cold acclimation and was characterized by the higher electric conductivity, compared to Aurora. These results indicated that higher winter hardiness of Aurotica was provided by different molecular mechanisms than free proline accumulation and changes in membrane that prevent electrolyte leakage after freezing stress. In the experiments of laboratory freezing of Aurora and Aurotica seedlings (crowns) and the following survival assessment, Aurotica demonstrated better survival. During the preparation for the freezing experiments, when seedlings were excavated from the soil after acclimation, a useful observation was made that Aurotica formed more developed (stronger) root system compared to Aurora when grown at low positive temperature (4°C). Based on this observation, the next root system characteristics were studied for Aurotica, Aurora and introgressive lines. For the root system study plants were grown in the paper boxes in laboratory and also in the field. Such root system characteristics as maximum root length, root number and root volume (determined as volume of liquid supplanted by the roots) were determined. Difference between Aurora and Aurotica was identified for root volume. It was shown that introgressive lines with higher correlation of root volume to the number of productive shoots were characterized by better field survival. Primers were developed to the coding sequence of the gene AGL21, which regulates root development via positive regulation of auxin accumulation in lateral root primordia. Aurora and Aurotica had polymorphic products with primers to second part of the gene. Introgressive lines were analyzed, and it was demonstrated that lines with AGL21 allele specific for Aurotica developed root system with greater volume at cold acclimation conditions, and were characterized by better winter field survival.
Автореферат диссертации на соискание ученой степени кандидата биологических наук по специальности 03.00.15 – генетика. – Институт клеточной биологии и генетической инженерии НАН Украины, Киев, 2017. В диссертации показано связь характеристик корневой системы с повышенной зимостойкостью для амфидиплоида Авротика (ААВВТТ) и интрогрессивных линий пшеницы с генетическим материалом генома Т вида Aegilops mutica. Созданные интрогрессивные линии, которые происходят от скрещивания сорта мягкой пшеницы Аврора и амфидиплоида Авротика, оценены с использованием микросателитного анализа относительно присутствия интрогрессий в составе хромосом 5 гомеологической группы. Показано присутствие "мозаических" хромосом, которые могли возникнуть в результате рекомбинации геномов D и Т. Определено, что накопление свободного пролина и изменения проницаемости мембраны не относятся к механизмам обеспечения высокой зимостойкости амфидиплоида Авротика. Установлено позитивную связь между объемом корневой системы Авротики и интрогрессивных линий и выживанием растений в полевых условиях зимой. Показано, что характерный для Авротики аллель гена AGL21, продукт которого регулирует синтез ауксина в корнях, встречается чаще у растений, которые перезимовали в поле, следовательно, может иметь связь с зимостойкостью растений.
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Keywords
інтрогресивні лінії, Amblyopyrum muticum, Triticum aestivum, зимостійкість, коренева система, транскрипційні фактори мікросателітні локуси, ауксин, автореферат дисертації, introgressive lines, Amblyopyrum muticum, Triticum aestivum, root system, winter hardiness, SSR, transcription factors, auxin, thesis abstract, интрогрессивные линии, Amblyopyrum muticum, Triticum aestivum, зимостойкость, корневая система, микросателитные локусы, транскрипционные факторы, ауксин, автореферат диссертации