Gel imager

SDS-PAGE analysis was performed by the method of Wang et al. (2022) (link) on 12 % resolving gel with 5 % stacking gel. Briefly, 5 % (w/v) SDS solution was added to the purified TLP sample, and the mixture was incubated in a water bath (85 °C, 1 h). The mixture was dispersed over 2 min at a speed of 4000 r/min using an oscillator (SHA-B, SHA-B, Liang You experimental instrument factory, Shanghai, China). Following this, the sample was centrifuged at 5000 × g for 5 min (4 °C). The resulting supernatant (15 uL) was subjected to the conditions of electrophoretic separation. A Mini Protean cell apparatus (Bio-Rad Laboratories, Hercules, CA, USA) was used to conduct these experiments. The gel was stained with 0.25 % (w/v) Coomassie Brilliant Blue R-250 and destained in 25 % (v/v) methanol solution containing 7.5 % (v/v) acetic acid. The gel was photographed following the processes of staining and destaining using a gel imager (GE Healthcare, UK).

For detection of OmpA, A. baylyi cells were grown in LB and adjusted to OD₆₀₀ of 1 in SDS loading buffer. Proteins from lysed cells were separated on Mini‐PROTEAN^® Precast Gels (Bio‐Rad) and transferred to nitrocellulose membrane for immune detection. Primary antibody against OmpA (generated by Genscript) was added at concentration of 0.25 µg/ml in PBS buffer with 5% milk containing 0.1% Tween 20 (PBST). Membrane was incubated for 1.5 h, followed by another 1.5 h incubation with PBST containing horseradish peroxidase‐labeled anti‐rabbit antibody (Jackson Lab). Peroxidase‐labeled conjugates were detected by LumiGLO Chemiluminescent Substrate (Seracare) on a Gel Imager (GE ImageQuant LAS 4000).