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Rational Design of Evolutionarily Stable Microbial Kill Switches
Molecular Cell ( IF 16.0 ) Pub Date : 2017-11-16 , DOI: 10.1016/j.molcel.2017.10.033
Finn Stirling , Lisa Bitzan , Samuel O’Keefe , Elizabeth Redfield , John W.K. Oliver , Jeffrey Way , Pamela A. Silver

The evolutionary stability of synthetic genetic circuits is key to both the understanding and application of genetic control elements. One useful but challenging situation is a switch between life and death depending on environment. Here are presented “essentializer” and “cryodeath” circuits, which act as kill switches in Escherichia coli. The essentializer element induces cell death upon the loss of a bi-stable cI/Cro memory switch. Cryodeath makes use of a cold-inducible promoter to express a toxin. We employ rational design and a toxin/antitoxin titering approach to produce and screen a small library of potential constructs, in order to select for constructs that are evolutionarily stable. Both kill switches were shown to maintain functionality in vitro for at least 140 generations. Additionally, cryodeath was shown to control the growth environment of a population, with an escape frequency of less than 1 in 105 after 10 days of growth in the mammalian gut.



中文翻译:

进化稳定的微生物杀灭开关的合理设计

合成遗传电路的进化稳定性是理解和应用遗传控制元件的关键。一个有用但具有挑战性的情况是生与死之间的切换取决于环境。这里介绍了“增效器”和“催眠”电路,它们在大肠杆菌中起着杀灭开关的作用。当失去双稳态cI / Cro记忆开关时,必需剂元素诱导细胞死亡。死泪利用冷诱导型启动子来表达毒素。我们采用合理的设计和毒素/抗毒素滴定法来生产和筛选潜在构建物的小型文库,以选择进化稳定的构建物。两种杀灭开关均显示在体外可维持功能至少有140代。此外,低温细菌已显示可控制种群的生长环境,在哺乳动物肠道中生长10天后,其逃逸频率不到十分之(5/1)。

更新日期:2017-11-16
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