Phage display library construction

Oligonucleotides were synthesized by GeneArt (Thermo Fisher Scientific) using TRIM technology. The oligonucleotides were incorporated into the pADL-10b phagemid (Antibody Design Labs) (fig. S23). Library construction was performed following previously published protocols (9). In brief, electrocompetent SS320 cells (Lucigen) were electroporated with library DNA. Bacteria were grown on plates containing agar with 2xYT medium (Sigma-Aldrich) supplemented with carbenicillin. After harvesting the bacteria, phages were produced by infecting mid-log phase bacteria with M13K07 helper phage (Antibody Design Labs); resuspending in 2xYT medium with carbenicillin, kanamycin, and isopropyl-β-d-thiogalactopyranoside (Thermo Fisher Scientific); and growing overnight at 30°C. Phages were precipitated on ice followed by centrifugation and resuspension. Library DNA was subjected to next-generation sequencing as previously described (57).