首页> 美国卫生研究院文献>Proceedings of the National Academy of Sciences of the United States of America >From the Cover: Adaptive evolution of genomically recoded Escherichia coli
【2h】

From the Cover: Adaptive evolution of genomically recoded Escherichia coli

机译:从封面:基因组编码的大肠杆菌的适应性进化

代理获取
本网站仅为用户提供外文OA文献查询和代理获取服务,本网站没有原文。下单后我们将采用程序或人工为您竭诚获取高质量的原文,但由于OA文献来源多样且变更频繁,仍可能出现获取不到、文献不完整或与标题不符等情况,如果获取不到我们将提供退款服务。请知悉。

摘要

Efforts are underway to construct several recoded genomes anticipated to exhibit multivirus resistance, enhanced nonstandard amino acid (nsAA) incorporation, and capability for synthetic biocontainment. Although our laboratory pioneered the first genomically recoded organism (Escherichia coli strain C321.∆A), its fitness is far lower than that of its nonrecoded ancestor, particularly in defined media. This fitness deficit severely limits its utility for nsAA-linked applications requiring defined media, such as live cell imaging, metabolic engineering, and industrial-scale protein production. Here, we report adaptive evolution of C321.∆A for more than 1,000 generations in independent replicate populations grown in glucose minimal media. Evolved recoded populations significantly exceeded the growth rates of both the ancestral C321.∆A and nonrecoded strains. We used next-generation sequencing to identify genes mutated in multiple independent populations, and we reconstructed individual alleles in ancestral strains via multiplex automatable genome engineering (MAGE) to quantify their effects on fitness. Several selective mutations occurred only in recoded evolved populations, some of which are associated with altering the translation apparatus in response to recoding, whereas others are not apparently associated with recoding, but instead correct for off-target mutations that occurred during initial genome engineering. This report demonstrates that laboratory evolution can be applied after engineering of recoded genomes to streamline fitness recovery compared with application of additional targeted engineering strategies that may introduce further unintended mutations. In doing so, we provide the most comprehensive insight to date into the physiology of the commonly used C321.∆A strain.
机译:正在努力构建几个预期具有多重病毒抗性,增强的非标准氨基酸(nsAA)掺入和合成生物遏制能力的重新编码的基因组。尽管我们的实验室率先开发了第一个基因组编码的生物(大肠杆菌C321.ΔA),但其适应性远低于未编码的祖先,特别是在确定的培养基中。这种适应性缺陷严重限制了其在需要确定培养基的nsAA连锁应用中的效用,例如活细胞成像,代谢工程和工业规模的蛋白质生产。在这里,我们报道了在葡萄糖最低培养基中生长的独立复制种群中,C321.ΔA的适应性进化超过了1000代。进化后的已编码种群大大超过了祖先C321.ΔA和未编码菌株的增长率。我们使用了下一代测序技术来鉴定在多个独立种群中突变的基因,并通过多重自动基因组工程(MAGE)重建了祖先菌株中的各个等位基因,以量化其对适应性的影响。几个选择性突变仅在重新编码的进化种群中发生,其中一些与响应于编码而改变翻译装置有关,而另一些与突变显然无关,而是纠正了在初始基因组工程期间发生的脱靶突变。该报告表明,与应用可能引入其他意外突变的其他靶向工程策略相比,可以对改造后的基因组进行工程改造后应用实验室进化,以简化适应度恢复。通过这样做,我们提供了迄今为止最常用的C321.∆A菌株的生理学最全面的见解。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
代理获取

客服邮箱:kefu@zhangqiaokeyan.com

京公网安备:11010802029741号 ICP备案号:京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有
  • 客服微信

  • 服务号