Larger fungi are of great value in natural ecosystems as decomposers of organic substances and mycorrhiza-forming organisms. Many species of larger fungi are known to be valuable sources of food, and of stimulating substances and drugs. Isolation and maintenance of cultures of larger fungi is an efficient way to conserve their biodiversity, but limited knowledge about these cultures and problems connected with their isolation make maintenance of many species impractical in laboratory conditions. The special culture collection of higher basidiomycetes at the M.G. Kholodny Institute of Botany maintains more than 600 strains of 200 species of larger Basidiomycetes. The authors have obtained pure cultures and have studied morphological features of the rare fungi including Anthurus archeri, Boletus aestivalis, Leucocoprinus bresadolae and Macrolepiota puellaris. Growth rate, and optimum and critical temperatures for growth were determined, and culture characters and their variability were described. Techniques have been proposed using nutrient media with stimulators of plant origin maintain viability of fungi from different ecological groups. New knowledge about morphogenesis in culture using light and electronic microscopy was obtained. A current project is to obtain cultures and create strain diversity of medicinal mushrooms. More than 70 species found to have medicinal features are maintained in the collection. Cultures of Lentinus edodes, Pleurotus ostreatus, Hericium erinaceum, Agrocybe aegerita, Grifola frondosa, Auricularia auricula, Auricularia polytricha and others were selected on the basis of a strategy of stepped screening. This research is a step to practical use in Ukraine of mushrooms long valued in oriental medicine. Biodiversity of valuable edible mushrooms is also represented in the collection, which is a very important source of the fruitbodies and biomass needed for the development of a mushroom growing industry in Ukraine. This research is supported by Ukrainian State Fund of Fundamental Research and the German Academic Exchange Service (DAAD).