Fig. 3: Continuous lytic cycling of phagemids by multiplicity tuning.

a, Change in optical density of E. coli cultures expressing T7 DNAP wild-type and variants (v2.4–v9) when infected with T7ΔDNAP over 3 hours at varying MOIs (0.01–10). Data are shown as mean ± s.d. of n = 3 independent experiments. b, Lysis kinetics of E. coli cultures expressing wild-type (dashed lines) or hypermutagenic (v9; solid lines) T7 DNAP at different MOIs (0–10). c, Schematic representation of proposed multiplicity tuning mechanism: after infection (step I), wild-type T7 DNAP enables efficient phage replication at a low MOI (step IIa), while error-prone variant v9 requires a higher MOI due to increased phage inactivation during replication (step IIb). d, Quantification of phagemid packaging efficiency across T7 DNAP variants, showing maintained library diversity despite reduced transduction rates in error-prone variants. Packaging efficiency is reported as CFU ml⁻¹ phage lysate. Data are shown as mean ± s.d. of n = 3 independent experiments. e, Representative 20-hour time course of a complete LySE cycle, showing distinct phases of bacterial growth and phage-mediated lysis initiated by simple mixing of phage lysates and cell cultures. Diagrams in c and e created in BioRender; Fredens, J. https://biorender.com/8aqhp97 (2026).