EXPERIMENTAL MODELS OF VIRAL ADAPTATION

病毒适应的实验模型

• 批准号: 2650507
• 负责人: JAMES J BULL
• 金额: $ 24.1万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-01-01 至 2001-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2650507
• 关键词: bacterial virus; chemical substitution; environmental adaptation; evolution; molecular biology; nucleic acid sequence; oligonucleotides; polymerase chain reaction; population genetics; virus genetics

DESCRIPTION: The goal of this study is to develop an approach to study the rules of viral adaptation at the molecular level that may inform us about how to develop anti-viral therapies. There are 3 specific aims to this proposal. (1) To examine the population genetics of viral populations over time. The data generated related to this aim will allow three hypotheses to be addressed. Is the pattern of substitution indicative of a series of selective sweeps? Are most of the fitness improvements due to a few substitutions with major effect and most of the substitutions will have very little effect? As the population reaches equilibrium will the level of variation decay to one sequence? (2) To explore the implications of parallel and convergent evolution during viral adaptation. Will parallel changes occur most of the time at sites with major as opposed to minor fitness benefits? Will related viruses adapt to the same selection with the same changes? (3) To examine the fitness cost of viral adaptation to multiple agents. Is there a fitness cost to adapting to a new selective agent? Can responses to multiple agents be predicted from responses to single agents? Is adaptation dependent on the prior history of adaptation? The PI proposes to use phage chemostats and adapt replicate populations to 43.5C. Samples will be taken each 6 hours for 3 days, then each 12 hours for 7 days. 96 samples will be taken each time point and assayed for a set of substitutions by oligonucleotide screening of predetermined regions (the set chosen from a pilot genome sequencing efforts). Fitness will be assayed (as the increase in phage per hour after infection of bacteria). These data will be used to address specific aim #1 (population genetics of viral adaptations over time) Six replicates of the first experiment will be undertaken (3 at a separate institution). These data will allow the PI to address specific aim #2 (the implications of convergent evolution during phage adaptation). One isolate from each replicate will be sequenced to determine screening sites. A second phase of research involves selecting against multiple agents. A series of agents, such as alcohol's, salts, etc. will be used as the selective agents. Selection will take place on a number of different media, as needed. Phage will be evolved that are resistant to a large number of agents, both separately and in combination. These data will allow the PI to address specific aim #3 (the fitness costs of adaptation to multiple agents).

描述：本研究的目的是开发一种方法来研究 病毒在分子水平上的适应规则可以告诉我们 如何开发抗病毒疗法。 对此有3个具体目标 提议。 (1) 检查病毒种群的种群遗传学 时间。 与该目标相关的数据将允许三个假设 得到解决。 替代模式是否表明一系列 选择性扫描？ 大多数健身改善是由于某些原因吗？ 具有重大影响的替换，并且大多数替换都会产生非常大的影响 影响不大？ 当人口达到平衡时， 变异衰减到一个序列？ (2) 探讨其含义 病毒适应过程中的平行进化和趋同进化。 将并行 大多数情况下，更改发生在重大而不是次要的站点上 健身的好处？ 相关病毒是否会适应与相同的选择？ 同样的变化？ (3) 检查病毒适应的适应度成本 多个代理。 适应新的选择是否有适应成本 代理人？ 可以根据对多个代理的响应来预测对多个代理的响应吗？ 单一代理人？ 适应是否依赖于之前的适应历史？ PI 建议使用噬菌体恒化器并调整复制群体以适应 43.5℃。 三天内每 6 小时取样一次，然后每 12 小时取样一次 7天。 每个时间点将采集 96 个样本并进行一组分析 通过寡核苷酸筛选预定区域（ 从试点基因组测序工作中选择的集合）。 将进行体能测试 （作为细菌感染后每小时噬菌体的增加）。 这些数据 将用于解决具体目标#1（病毒的群体遗传学） 随着时间的推移进行调整） 将进行第一个实验的六次重复（其中 3 次在单独的 机构）。 这些数据将使 PI 能够实现特定目标#2（ 噬菌体适应过程中趋同进化的影响）。 一种分离物 对每个重复的样品进行测序以确定筛选位点。 一个 研究的第二阶段涉及对多个代理进行选择。 一个 采用醇类、盐类等系列试剂作为 选择剂。 选择将在许多不同的媒体上进行， 根据需要。 噬菌体将进化出对大量噬菌体具有抵抗力的噬菌体。 代理，既可以单独使用，也可以组合使用。 这些数据将使 PI 能够 解决具体目标#3（适应多种环境的适应成本） 代理）。