Структурно-функциональная организация рибосомных повторов человека
Диссертация
Для клеток млекопитающих (крысы и мыши) показано, что РГ, с точки зрения конформации, делятся на две фракции (Conconi et al., 1989): «открытую» и «закрытую». В «открытой» конформации (ТО рДНК в целом повышенно доступна действию различных агентов) находятся активные и потенциальноактивные копии РГ. В литературе не приводится более конкретных данных об организации копий «открытой» («закрытой… Читать ещё >
Содержание
- ГЛАВА 1. НЕРАДИОАКТИВНОМЕЧЕНЫЕ ДНК-ЗОНДЫ- ОСОБЕННОСТИ ФРАГМЕНТАЦИИ рДНК
- 1. 1. Нерадиоактивномеченые ДНК- зонды для гибридизации
- 1. 1. 1. Обзор литературы
- 1. 1. 2. Задачи исследования
- 1. 1. 3. Синтез и свойства фотоактивируемых арилазидов
- 1. 1. 4. Получение и свойства ДНК-зондов
- 1. 1. 5. Использование фотобиотинилированных ДНК-зондов
- 1. 1. 5. 1. Дот -, блот — и in situ — гибридизация
- 1. 1. 5. 2. Количественные методы анализа последовательностей ДНК
- 1. 1. 5. 3. Зонды на транскрибируемую область рибосомного повтора
- 1. 1. Нерадиоактивномеченые ДНК- зонды для гибридизации
- 1. 2. Особенность фрагментации ТО рДНК при действии реагентов, индуцирующих в ДНК однонитевые разрывы
- 1. 2. 1. Анализ длины фрагментов рДНК в образцах ДНК, выделяемой из различных тканей и биожидкостей
- 1. 2. 2. Устойчивость ТО рДНК к фрагментации при внесениии в ДНК однитевых разрывов
- 1. 2. 3. Практическое применение свойства устойчивости ТО рДНК к парным разрывам
- 1. 2. 3. 1. Детекция фрагментов рДНК в сыворотке крови человека в норме и при патологии
- 2. 1. Обзор литературы
- 2. 1. 1. Общее число копий РГ в геноме человека
- 2. 1. 2. Количество копий рДНК в ЯОР хромосом человека
- 2. 1. 3. Определение количества активных копий рДНК
- 2. 1. 3. 1. Анализ сателлитных ассоциаций, Ag-окраска хромосом
- 2. 1. 3. 2. Метод непрямой иммунофлуоресценции
- 2. 1. 3. 3. Потенциально активные копии РГ млекопитающих
- 2. 1. 4. Задачи исследования
- 2. 2. Число копйй рДНК в геноме человека
- 2. 2. 1. Объекты исследования
- 2. 2. 2. Метод определения числа копий РГ в геноме
- 2. 2. 2. 1. Выделение ДНК из клеток
- 2. 2. 2. 2. Измерение концентрации ДНК и ее фрагментация
- 2. 2. 2. 3. Контрольные последовательности- калибровочные кривые
- 2. 2. 2. 4. Условия гибридизации- характеристика зондов
- 2. 2. 2. 5. Дополнительные контрольные эксперименты
- 2. 2. 2. 6. Число коп ий РГ в геноме человека
- 2. 3. 1. Объекты исследования
- 2. 3. 2. Число активных копий РГ в стимулированных лимфоцитах
- 2. 3. 3. Число неактивных копий РГ, содержащихся в активных и неактивных ЯОР хромосом
- 2. 4. 1. Объекты исследования
- 2. 4. 2. Определение двух фракций РГ
- 2. 4. 2. 1. Кинетика гидролиза ядер лимфоцитов человека рестриктазой Rsal
- 2. 4. 2. 2. Количественные соотношения между копиями РГ, находящимися в «открытой «и «закрытой «конформации
- 3. 1. Обзор литературы
- 3. 1. 1. Общие сведения о метилировании ДНК в клетках млекопитающих
- 3. 1. 2. Последствия метилирования ДНК в клетке
- 3. 1. 3. Методы изучения метилирования геномной ДНК
- 3. 1. 4. МетилированиерДНКмлекопитающих
- 3. 1. 5. Связь метилирования и транскрипции рибосомных генов
- 3. 1. 6. Задачи исследования
- 3. 2. Метилирование ТО рДНК лейкоцитов человека
- 3. 2. 1. Объекты исследования
- 3. 2. 2. Методы изучения метилирования рДНК
- 3. 2. 3. Анализ метилирования ТО рДНК человека с помощью гидролиза рестриктазой Hpall
- 3. 2. 4. Гидролиз ТО рДНК человека рестриктазой Bsp
- 3. 2. 5. Изучение метилирования ТО рДНК с помощью рестриктазы Hinfl
- 3. 2. 6. Количественная оценка метилирования тотальной ДНК и рДНК
- 3. 2. 7. Сравнение результатов цитогенетического анализа РГ и данных о метилировании ТО рДНК
- 3. 3. Метилирование ТО рДНК в различных типах клеток
- 3. 3. 1. Метилирование рДНК в лимфоцитах и лейкоцитах
- 3. 3. 2. Метилирование ТО рДНК в сперматозоидах человека
- 3. 3. 3. Метилирования рДНК в различных тканях
- 3. 4. Обсуждение результатов и
- 3. 4. 1. Вариабельность метилирования ТО рДНК человека
- 3. 4. 2. Вариабельность метилирования копий рДНК одного генома
- 3. 4. 3. Сопоставление количеств неактивных и метилированных копий
- 3. 4. 4. Метилирование ТОрДНК в различных типах клеток человека
- 3. 4. 5. Возможный механизм образования паттерна метилирования рДНК
- 4. 1. Обзор литературы
- 4. 1. 1. Локализация РГ в структурах ядрышка
- 4. 1. 2. Расположение транскрибируемых копий РГ
- 4. 1. 3. Взаимодействие рДНК с ядерным матриксом
- 4. 1. 4. Конформация потенциальноактивных и неактивных копий РГ
- 4. 1. 5. Задачи исследования
- 4. 2. Доступность различных участков ТО РГ в клетках и ядрах человека действию эндонуклеаз и арилазида
- 4. 2. 1. Объекты исследования
- 4. 2. 2. Rsal — гидролиз ТО рДНК в изолированных ядрах лимфоцитов
- 4. 2. 3. Rsal — гидролиз ТО рДНК в изолированных ядрах фибробластов и клеток лимфобластоидной линии
- 4. 2. 4. Гидролиз ТОрДНКядер лимфоцитов ДНКазой
- 4. 2. 5. Модификация арилазидом ТО рДНК в лимфоцитах и изолированных ядрах лимфоцитов
- 4. 3. Прочносвязанные с рДНК белки РП человека
- 4. 3. 1. Обнаружение прочносвязанных белков
- 4. 3. 2. Прочносвязанные с рДНК белки в изолированных ядрах, нуклеоиде и нуклеопротеиде
- 4. 3. 3. Выделение и частичная характеристика белков, экранирующих R-caum
- 4. 3. 4. Количество копий РГ с экранированным Rsal- сайтом 10 701 при различных состояниях клетки
- 4. 4. Расположение метилированных копий РГ в ядрах лимфоцитов
- 4. 5. Обсуждение результатов
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