PDF(Pubmed)
Abstract：
Sophisticated gene circuits built by synthetic biology can enable bacteria to sense their environment and respond predictably. Engineered biosensing bacteria outfitted with such circuits can potentially probe the human gut microbiome to prevent, diagnose, or treat disease. To provide robust biocontainment for engineered bacteria, we devised a Cas9-assisted auxotrophic biocontainment system combining thymidine auxotrophy, an Engineered Riboregulator (ER) for controlled gene expression, and a CRISPR Device (CD). The CD prevents the engineered bacteria from acquiring thyA via horizontal gene transfer, which would disrupt the biocontainment system, and inhibits the spread of genetic elements by killing bacteria harboring the gene cassette. This system tunably controlled gene expression in the human gut commensal bacterium Bacteroides thetaiotaomicron, prevented escape from thymidine auxotrophy, and blocked transgene dissemination. These capabilities were validated in vitro and in vivo. This biocontainment system exemplifies a powerful strategy for bringing genetically engineered microorganisms safely into biomedicine.
摘要：
合成生物学构建的复杂基因回路可以使细菌感知其环境并做出可预测的反应。配备此类电路的工程生物传感细菌可以潜在地探测人类肠道微生物群以防止，诊断,或治疗疾病。为工程细菌提供强大的生物防护，我们设计了一个Cas9辅助的营养缺陷型生物保护系统，结合了胸苷营养缺陷型，用于控制基因表达的工程化核糖调节剂（ER），和CRISPR设备(CD)。CD阻止工程化细菌通过水平基因转移获得thyA，这会破坏生物安全壳系统，并通过杀死携带基因盒的细菌来抑制遗传因子的传播。该系统可调节地控制人类肠道共生细菌中的基因表达，防止逃避胸苷营养缺陷型，并阻止转基因传播。这些能力在体外和体内得到验证。这种生物保护系统体现了将基因工程微生物安全地带入生物医学的强大策略。
