A method for isolating high purity and quantity RNA from Agaricus bisporus which is rich in proteins, carbohydrate, fiber and secondary metabolites, is described. RNA was extracted from mycelium, primordia, sporophores at two development stages and two post-harvest storage stages as well as from pileipellis, inner cap, gill and stipe of the mature sporophore. The A(260)/A(230) and A(260)/A(280) ratios of isolated RNA from fruiting bodies were both similar to 2 and the yield was about 200 mu g/g fresh wt (FW). The yield of RNA from mycelium was approx. 100 mu g/g FW. High quality RNA was also extracted from fruiting body tissues of Lentinus edodes, Pleurotus ostreatus, Flammulina velutipes and Pleurotus eryngii with yields from 130 to 225 mu g/g FW. RNA extracted from all samples was intact, as demonstrated by gel electrophoresis and was suitable for downstream molecular applications, including RT-PCR and qPCR.