Versatile and iterative prokaryotic genome engineering through CRISPR-Transposon phage particles. Escherichia coli

The engineering of microbial genomes is key for the generation of microbial chassis tailored towards human innovation and biotechnological applications. However, prokaryotic genome engineering has been greatly hindered by the innate feasibility, scales, and ease of delivery and integration of genetic payloads in non-model microbial organisms. Here we address these challenges through a Phage Assisted CAST Integration (PAINT) system, and its iterative counterpart (iPAINT), which synergistically couples phage packaging and CAST transposition machinery to achieve reliable, streamlined, and highly efficient targeted genome insertions in a wide array of microbial species.