Streptomycin-Resistant E. coli Strain S-17 Characterization

streptomycin-resistant green fluorescent protein (GFP)-expressing E. coli strain S-17 (S-17-GFP) was generated as described previously^{75 (link)}. S-17-GFP were cultured in Luria-Bertani medium (Nacalai Tesque) with 50 μg/mL streptomycin (Wako) overnight, then bacteria were transferred into LB medium with 1 mM isopropyl β-D-1-thiogalactopyranoside (Wako) and incubated at 37 °C for an hour. After incubation, bacteria were resuspended in 800 μL of 4% paraformaldehyde at 4 °C for 3 h. Then bacteria were resuspended into PBS and measured OD 600 by spectrophotometer. A total of 1 × 10⁷ cells (assuming OD 600 1.0 = 5 × 10⁸ cell/mL) bacteria were transferred to a new tube and resuspended and incubated with 10 μg rGP2 or PBS at 4 °C overnight. GP2-bound bacteria were detected by using a combination of anti-mGP2 antibody diluted 1:100 and PE-conjugated anti-rat IgG2a antibody diluted 1:100. Flow cytometry analysis was performed with a FACSCanto II instrument or ATTUNE Next. Gating strategies for the bacterial flow cytometry were shown in Supplementary Fig. 18.