This study was conducted to investigate optimum cultural conditions for Amanita hemibapha isolated in Korea and its hyphae characteristics. Micrographs shows the presence of clamp connection. A. hemibapha grows as mycelial form(M-phase) 2-4 μm and yeast-like form(Y-phase) 7-8 μm. The fungal spores were broadly elliptical and papillate, 8-11 × 6-9 μm in size. The nucleotide sequence analysis of the ITS of nuclear ribosomal DNA from sporocarps and in-vitro-grown mycelium supported the fungal species is Amanita hemibapha. A. hemibapha showed sequence similarity in the ITS rDNA with A. caesarea(97.5) and A. jacksonii(98.5%) which are morphologically similar species to A. hemibapha. The optimal pH and temperature for mycelial growth of A. hemibapha were pH 6.0 and 28 oC, respectively. The fungal species showed best growth in SYP and GYS medium. A. hemibapha grew well with mannitol and glucose as carbon sources and peptone as a nitrogen source.