Isolation of the MLS-type Flagellar Apparatus from the an-1 mutant of Fern Lygodium japonicum and Its Preliminary Immunological Analysis

The basal apparatuses preserving the multilayered structure (MLS) were isolated from sperm of the an-1 strain of fern Lygodium japonicum by detergent extraction and DNase digestion, followed by Percoll discontinuous density centrifugation. Isolated basal apparatuses retained the spiral shape as that of the intact sperm and was consisted of the broad flagellar root (spline), MLS, crest, basal bodies and electron-dense material embedding the basal bodies (embedding material). In the isolated basal apparatus fine filaments were found between the basal bodies and the spline by electron microscopy. Three monoclonal antibodies (mAb h1c, 1C7 and 13D10) raised against the purified basal apparatuses were used to explore novel proteins in the MLS-type basal apparatus. A 42 kD protein recognized by mAb h1c was localized in the crest by immunogold electron microscopy. mAb 1C7 reacted with a 41 kD protein and 13D10 with a 43 kD in western blotting, though both antibodies uniformly decorated the embedding material by immunoelectron microscopy.