Bacteroides thetaiotaomicron is a human-associated bacterium that holds promise for delivery of therapies in the gut microbiome1. Therapeutic bacteria would benefit from the ability to turn on different programs of gene expression in response to conditions inside and outside of the gut; however, the availability of regulatory parts, and methods to combine them, have been limited in B. thetaiotaomicron2-5. We report implementation of Cello circuit design automation software6 for this species. First, we characterize a set of genome-integrated NOT/NOR gates based on single guide RNAs (CRISPR-dCas9) to inform a Bt user constraint file (UCF) for Cello. Then, logic circuits are designed to integrate sensors that respond to bile acid and anhydrotetracycline (aTc), including one created to distinguish between environments associated with bioproduction, the human gut, and after release. This circuit was found to be stable under laboratory conditions for at least 12 days and to function in bacteria associated with a primary colonic epithelial monolayer in an in vitro human gut model system.

中文翻译：

---

肠道常驻物种 Bacteroides thetaiotaomicron 的遗传回路设计自动化。

Bacteroides thetaiotaomicron 是一种与人类相关的细菌，有望在肠道微生物组中提供治疗。治疗性细菌将受益于根据肠道内外条件启动不同基因表达程序的能力；然而，在 B. thetaiotaomicron2-5 中，监管部分的可用性和组合它们的方法受到限制。我们报告了该物种的 Cello 电路设计自动化软件 6 的实施情况。首先，我们描述了一组基于单向导 RNA (CRISPR-dCas9) 的基因组整合的 NOT/NOR 门，以通知 Cello 的 Bt 用户约束文件 (UCF)。然后，逻辑电路被设计成集成对胆汁酸和无水四环素 (aTc) 有反应的传感器，包括一个用来区分与生物生产相关的环境的传感器，人体肠道，释放后。发现该回路在实验室条件下稳定至少 12 天，并且在体外人类肠道模型系统中与原代结肠上皮单层相关的细菌中发挥作用。