Construction of Sniper1 Mutant Libraries

Sniper1 mutant libraries were constructed using three independent protocols for mutagenesis from XL1-red competent cells (Agilent), Genemorph II (Agilent) and Diversify polymerase chain reaction (PCR) random mutagenesis (Clontech) kits. All reaction conditions have been described previously^{7 (link)}. The assembled libraries were transformed into Endura electrocompetent cells (Lucigen) and incubated on LB plates containing chloramphenicol (12.5 μg ml⁻¹) at 37 °C overnight. A total of 3 × 10⁶ colonies were obtained for each library, resulting in an overall library complexity of 10⁷. Pooled library plasmids were purified using a midi prep kit (NucleoBond Xtra Midi EF; Macherey-Nagel).