Fig. 2.
TfcP, minor pilins, and PilY1.1 of cluster_1 are important for T4aPdM and T4aP formation. (A) Assay for T4aPdM. WTΔ2Δ3 and strains with deletions of individual cluster_1 genes, and the corresponding complementation strains were spotted on 0.5% agar supplemented with 0.5% CTT and imaged after 24 h. (Scale bar, 1 mm.) (B) Shearing assay for T4aP formation. (Top) T4aP sheared off from ∼15 mg cells grown on 1.5% agar supplemented with 1.0% CTT were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and probed with α-PilA antibodies. (Middle) A total of 40 µg of total protein from total cell extracts separated by SDS-PAGE and probed with α-PilA antibodies and (Lower) after stripping, with α-LonD antibodies as a loading control. (C) Shearing assay for T4aP formation in retraction-deficient strains. T4aP formation was assayed as in B. In lanes labeled *, fivefold less total protein was loaded.