[3S02a] 05. Genome Editing Technology and its Applications - Part 2

3:25 PM - 3:40 PM JST(6:25 AM - 6:40 AM UTC)

[3GET-12-KL] Novel nuclease enables targeted, continuous evolution of bacteriophages and host range expansion

○Aung Khine Linn^1,2, Yulan Liu^1,2, Julius Fredens^1,2 (1. National University of Singapore (Singapore), 2. National Centre for Engineering Biology (Singapore))

Keywords:

continuous evolution,phage host-range engineering,RNA-guided nickase,bacterial resistance,phage therapy

Phage therapy is constrained by the narrow host range of phages and the rapid adaption of membrane proteins and lipopolysaccharides (LPS) on the bacteria. To address these challenges, we adapted the EvolvR system from E. coli, where a fusion of nCas9 and an error-prone polymerase enables targeted, continuous mutagenesis of phage genomes in Escherichia coli. While nCas9 targets the phage T4 genome poorly, we identified a novel RNA-guided nickase to be highly efficient, and we successfully evolve T4 phage tail fibers for host-range expansion. Our continuous evolution platform supports modular targeting of distinct phage structural components and generates time-resolved libraries that can be screened against diverse host backgrounds. It offers a generalizable approach for phage evolution and it is readily extensible to other phages and pathogens, with potential applications in overcoming narrow host range and resistance barriers in phage therapy.