This study was carried out to investigate the promoting effect of wood flour on the mycelial growth of Lentinus lepideus and Lentinus edodes. To determine the optimal culture condition, we first examined the tissue origin of pine flour (Pinus densiflora) including needle, bark, root and xylem. Only the xylem-derived flour increased mycelial growth compared to no treatment control. The addition of the xylem flour (5 g/l) showed the highest increase and the glucose level in the basal medium was best at 10 g/l. The smaller particle size of the xylem flour showed the positive effect on mycelial growth; two-fold increase when supplemented with flour of which particle size is less than 106 µm in diameter compared to 425 µm. The addition of the xylem flour continuously increased the mycelial production for 25 days while mycelia stopped growing within 15 days without the xylem flour. In addition, when woody flour obtained from the different tree species was applied to L. edodes mycelial culture, all treatments accelerated mycelial production compared to the control. Based on all results described above, we conclude that the supplementation of woody flour to culture medium may be an another promising way to increase mycelial production of economically important fungi.