Abstract: Five strains of fungal endophytes, named YX1-6, YX1-7, QJ3-6, QJ3-7, QJ3-8, were isolated from four spieces of Boletus specious Frost, Boletus aereus, Leccinum crocipodium, Boletus edulis which were collected in Yuxi and Qujing areas of Yunnan Province.The morphyological characters of the strains were observed, and their ITS sequences were amplified and sequenced.Phylogenetic tree was constructed by the obtained molecular data.The results showed that YX1-6 was Trametes hirsuta., YX1-7 was Chaetomium aureum., QJ3-6 was Candida sp., QJ3-7 was Mortierella sp., QJ3- 8 was Eurotium sp.. The secondary structure of ITS rRNA in these five strains predicted with the softwares of DNA STAR, RNAVIZ2.0. and MEGA5.1 indicated that fungal endophytes from Boletaceae were different and diversified.