The Isolation of Nuclei from the Filamentous Fungus Aspergillus nidulans

Abstract

A method was developed for isolating nuclei from the filamentous fungus A. nidulans. In this procedure, the mycelia from 14-16 h spore-derived cultures of A. nidulans NRI, a stable diploid strain, were frozen with liquid N and homogenized in a Waring blender. After homogenization, the mycelia were warmed to 4.degree. C and the nuclei were purified from the homogenate by differential centrifugation followed by sedimentation through 2.1 M-sucrose. The final nuclear yield was 15-20%, based on DNA estimations. The purified nuclear pellet was free of whole cells. The morphology of the isolated nuclei resembled that of the in situ nuclei; they contained a nucleolus, chromatin, and had a surrounding double membrane. The purified nuclei were characterized by a DNA:RNA:protein ratio of 1:2.8:8.7.