Division of Infrastructure and Environment, School of Engineering, University of Glasgow, Rankine Building, Level 5, Glasgow, G12 8QQ, UK.
Microb Cell Fact. 2018 Jan 22;17(1):8. doi: 10.1186/s12934-018-0858-2.
Synthetic organism-based biotechnologies are increasingly being proposed for environmental applications, such as in situ sensing. Typically, the novel function of these organisms is delivered by compiling genetic fragments in the genome of a chassis organism. To behave predictably, these chassis are designed with reduced genomes that minimize biological complexity. However, in these proposed applications it is expected that even when contained within a device, organisms will be exposed to fluctuating, often stressful, conditions and it is not clear whether their genomes will retain stability.
Here we employed a chemostat design which enabled us to maintained two strains of E. coli K12 under sustained starvation stress: first the reduced genome synthetic biology chassis MDS42 and then, the control parent strain MG1655. We estimated mutation rates and utilised them as indicators of an increase in genome instability. We show that within 24 h the spontaneous mutation rate had increased similarly in both strains, destabilizing the genomes. High rates were maintained for the duration of the experiment. Growth rates of a cohort of randomly sampled mutants from both strains were utilized as a proxy for emerging phenotypic, and by association genetic variation. Mutant growth rates were consistently less than rates in non-mutants, an indicator of reduced fitness and the presence of mildly deleterious mutations in both the strains. In addition, the effect of these mutations on the populations as a whole varied by strain.
Overall, this study shows that genome reductions in the MDS42 did not stabilize the chassis under metabolic stress. Over time, this could compromise the effectiveness of synthetic organisms built on chassis in environmental applications.
基于合成生物体的生物技术越来越多地被提议用于环境应用，例如原位传感。通常，这些生物体的新功能是通过在底盘生物体的基因组中编译遗传片段来实现的。为了可预测地表现，这些底盘被设计成具有最小化生物复杂性的简化基因组。然而，在这些提议的应用中，即使被包含在设备内，生物体也可能会暴露于波动的、通常是应激的条件下，目前尚不清楚它们的基因组是否会保持稳定。
在这里，我们采用了恒化器设计，使我们能够在持续饥饿应激下维持两种大肠杆菌 K12 菌株：首先是简化基因组合成生物学底盘 MDS42，然后是对照亲本菌株 MG1655。我们估计了突变率，并将其用作基因组不稳定性增加的指标。我们表明，在 24 小时内，两种菌株的自发突变率都相似地增加，使基因组不稳定。在实验过程中，高突变率得以维持。我们利用两种菌株中随机抽样的突变体的群体生长率作为新兴表型的替代指标，以及遗传变异的关联。突变体的生长率始终低于非突变体的生长率，这表明两种菌株的适应性都降低，并且存在轻度有害突变。此外，这些突变对整个种群的影响因菌株而异。
总的来说，这项研究表明，在代谢应激下，MDS42 中的基因组减少并没有使底盘稳定。随着时间的推移，这可能会影响到基于底盘的合成生物体在环境应用中的有效性。
