Получение рекомбинантных антигенов вируса ящура О/Тайвань/99, перспективных для разработки противоящурных вакцин, с использованием бактерий и растений
Диссертация
Основной причиной распространения ящура является международная торговля домашними животными. Вспышки заболевания происходят во всех странах мира, где в сельском хозяйстве используется крупный рогатый скот. Болезни подвержены все домашние парнокопытные животные. Наиболее восприимчив к заболеванию ящуром крупный рогатый скот, менее восприимчивы свиньи, овцы, козы, более 70 видов диких животных… Читать ещё >
Содержание
- ОПРЕДЕЛЕНИЯ, ОБОЗНАЧЕНИЯ И СОКРАЩЕНИЯ
- ЦЕЛЬ И ЗАДАЧИ РАБОТЫ
- ОБЗОР ЛИТЕРАТУРЫ
- 1. Вирус ящура
- 2. Вакцины против ящура
- 2. 1. Традиционная вакцина
- 2. 2. ДНК-вакцины
- 2. 3. Вакцины на основе ослабленных вирусов
- 2. 4. Пустые вирусные капсиды как основа для вакцины
- 2. 5. Субъединичные вакцины
- 3. Антигенные сайты вируса ящура
- 4. Гетерологичные системы экспрессии белков
- 4. 1. Бактериальная система экспрессии
- 4. 2. Растительная система экспрессии
- 4. 2. 1. Трансгенные растения как система получения белков
- 4. 2. 2. Получение белков в хлоропластах
- 4. 2. 3. Транзиентная экспрессия белков в растениях
- 1. 1. Экспрессия предшественника капсидных белков Р1−2А в растениях N. Ьеп1кат1апа
- 1. 2. Коэкспрессия предшественника Р1−2А и протеазы ЗСР'° в N. benthamiana
- 1. 3. Экспрессия индивидуальных капсидных белков вируса ящура в растениях N. Benthamiana
- 2. 1. Получение белка CBD-PE в клетках Е. col
- 2. 2. Получение полиэпитопного белка НРЕ в Е. col
- 2. 3. Получение белка НРЕ в растениях N. benthamiana
- 2. 4. Схема получения полиэпитопного белка НРЕ из клеток бактерий и растений
- 2. 5. Оценка иммуногенности НРЕ белка
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- Определение протективного действия полученного в растительной системе экспрессии рекомбинантного полиэпитопного белка вируса ящура на морских свинках.1. Тестируемый препарат
- Препарат кандидатной вакцины ГПЭ растворен в 10 мл буфера A4 (10 мМ Tris-HCl pH 8.0- 4M мочевина) в концентрации 0,8 мг/мл. Визуально кандидатная вакцина представляет собой прозрачную жидкость. Условия хранения температура минус 20 °C.
- Способ введения препарата и доза
- Четвертая группа из 8 морских свинок, которым белок не вводили, была контрольной. Морских свинок иммунизировали однократно.
- Заражение морских свинок вирусом ящура
- Для оценки достоверности полученных результатов иммунизации использовали биномиальный тест с уровнем значимости (а) равным 0,005.1. РЕЗУЛЬТАТЫ
- Однократная иммунизация морских свинок выделенным из растений препаратом рекомбинантного полиэпитопного белка вируса ящура обеспечивает защиту животных от заражения вирусом ящура О/Тайвань/99 при использовании дозы препарата 120 и 350 мкг.
- Тестирование протективного действия полученного в бактериальной системе экспрессии рекомбинантного полиэпитопного белка вируса ящура на морских свинках.1. Тестируемый препарат
- Препарат кандидатной вакцины ГПЭ растворен в 5 мл буфера А4 (10 мМ Tris-HCl рН 8.0- 4 М мочевина) в концентрации 1,4 мг/мл. Визуально кандидатная вакцина представляет собой прозрачную жидкость. Условия хранения температура минус 20 °C.
- Способ введения препарата и доза
- Четвертая группа из 8 морских свинок, которым белок не вводили, была контрольной. Морских свинок иммунизировали однократно.
- Заражение люрских свинок вирусом ящура
- Для оценки достоверности полученных результатов иммунизации использовали биномиальный тест с уровнем значимости (а) равным 0,005.1. РЕЗУЛЬТАТЫ
- ГД5о -доза, вызывающая гибель 50% чувствительных клеток in vitro7