Escherichia coli is one of the most important microbial cell factories, but infection by bacteriophages in the environment may have a huge impact on its application in industrial production. Here, we developed a mobile CRISPR-Cas9 based anti-phage system for bacteriophages defense in E. coli. Two conjugative plasmids pGM1 (phosphoglucomutase 1) and pGM2 carrying one and two guide RNAs, respectively, were designed to defend against a filamentous phage. The results showed that the pGM1 and pGM2 could decrease the phage infection rate to 1.6% and 0.2% respectively in infected cells. For preventing phage infection in E. coli, the pGM2 decreased the phage infection rate to 0.1 %, while pGM1 failed to block phage infection. Sequence verification revealed that point mutations in protospacer or protospacer adjacent motif sequences of the phage genome caused loss of the defense function. These results support the potential application of MCBAS in E. coli cell factories to defend against phage infections.

大肠杆菌是最重要的微生物细胞工厂之一，但环境中的噬菌体感染可能对其在工业生产中的应用产生巨大影响。在这里，我们开发了一种基于 CRISPR-Cas9 的移动抗噬菌体系统，用于大肠杆菌中的噬菌体防御。两个结合质粒 pGM1（磷酸葡萄糖变位酶 1）和 pGM2 分别携带一个和两个引导 RNA，被设计用于防御丝状噬菌体。结果表明，pGM1和pGM2可以将感染细胞中的噬菌体感染率分别降低至1.6%和0.2%。用于防止大肠杆菌中的噬菌体感染, pGM2 将噬菌体感染率降低至 0.1%, 而 pGM1 未能阻断噬菌体感染。序列验证表明，噬菌体基因组的原型间隔区或原型间隔区相邻基序序列中的点突变导致防御功能丧失。这些结果支持 MCBAS 在大肠杆菌细胞工厂中的潜在应用，以防御噬菌体感染。