FIG. 3.
Biochemical fractionation of WalKSpn-(C)-FLAG, WalRSpn-(C)-L-FLAG3, and WalJSpn-(C)-L-FLAG3 in an exponentially growing unencapsulated derivative of strain D39. Fractionation and Western immunoblot assays using anti-FLAG antibody were performed as described in Materials and Methods. Total, supernatant (S) and pellet (P) of starting cultures. Wash, supernatant (S) and pellet (P) of cells washed with sucrose-containing buffer. Washed pellets were further separated into solubilized cell wall (CW) and protoplast (Prot) fractions. Lysed protoplasts were separated into cytoplasmic (Cyto) and membrane (Memb) fractions. The image shown is a semiquantitative autoradiogram. Quantitation in the text is based on three independent biological replicate experiments in which band luminescence was determined using an IVIS imaging system. See text for additional details.