P8, which was from Hm3-8, produced a 454-bp DNA fragment only for Hm1-1, Hm1-6, Hm2-10, and Hm3-8. The primer sets P1-P5 and P7 produced DNA bands with corresponding sizes from all H. marmoreus strains and presented no strain specificity (only P3 data are shown in Fig. 2a).

P1, P4, and P7 were polymorphic. The primer sets P6 and P8 could be employed for the specific detection of Hm1-1-related strains, while P9 and P10 were specific for Hm3-10. The specificities of the selected primer sets were challenged with the hybrid strains Hm15-3, Hm15-4, buy SRT1720 Hm15-5, Hm16-1, Hm16-2, and Hm17-5 (Fig. 2b). The P6 marker appeared only for Hm1-1, whereas the P8 marker appeared for most hybrid strains except Hm16-1 and the wild Hm3-10. This is interesting because the P6 marker showed broader specificity than the P8 marker in the identification of strains. The P9 marker appeared on Hm16-1, Hm16-2, Hm17-5, and Hm3-10 and the P10 marker appeared on Hm15-3, Hm15-4, Hm16-1, Hm16-2, and Hm3-10. The hybrid strain Hm15-3 was the only strain that did not contain either the P9 or the P10 marker. Development of new strains and verification techniques are some of the major issues in mushroom technology. In this study, we crossed a commercial strain of H. marmoreus and a wild strain of H. marmoreus by monokaryotic mycelial mating. The wild strain (Hm3-10) showed distinct morphological and cultivation characteristics.

Cultivated H. marmoreus PAK6 strains GSK-3 inhibitor review originated largely from Japan, where this mushroom is the second most cultivated mushroom. Most of them were raised from a few Japanese parental strains and thus are closely related to each other. Dendrogram analysis based on RAPD demonstrates that cultivated strains can be categorized in two groups (Fig. 1b) and the genetic distance between the groups is closer than that to Hm3-10.

Uniqueness of Hm3-10 was further evidenced by the mating experiment. Mating frequency between the commercial Hm1-1 and the wild Hm3-10 strain was 85.8%, which is unusually high for tetrapolar mating, indicating allelic diversification of the mating-type genes in the Korean strains. Similar results were reported in the mating of P. tuberregium from different geographic origins (Isikhuemhen et al., 2000). RAPD is not, in general, a good method for identification and classification of fungi because of limitations in reproducibility. However, it can be a simple and powerful tool when it is used to make comparisons within a set of samples. It is also a useful tool to generate SCAR markers (Weber et al., 2002; Tanaka et al., 2004). In this work, the primer sets derived from distinct RAPD bands were successfully employed to discriminate specific strains. Our results showed that PCR reactions with the primer sets yielded strain-specific DNA bands, indicating that our strategy to develop SCAR marker is a reasonable approach.