Morchella importuna is a worldwide distributed edible mushroom with high ecological and economic values, but the molecular and genetic research about this mushroom has been hindered due to lack of an efficient transformation method. Here, we report for the first time the successful transformation of M. importuna by using a hypervirulent Agrobacterium tumefaciens strain bearing the constructed binary plasmid p1391-U-GUS. The selectable markers used were the genes for hygromycin resistance under the control of the polyubquitin promoter from M. importuna. The reporter genes were those for enhanced green fluorescent protein (EGFP) and the β-Glucuronidase (GUS) under the control of glyceraldehyde-3-phosphate dehydrogenase promoter and polyubquitin promoter respectively. The presence of the reporter gene EGFP in the transformants was confirmed by the fluorescence and confocal microscope and molecular analysis and that of the reporter gene GUS was verified by enzyme activity and molecular analysis. The analysis results of both reporter genes indicated that Agrobacterium-mediated transformation was successfully performed in M. importuna.

Our previous study found that LeDnaJ07 RNAi decreased Lentinula edodes resistance to heat stress and Trichoderma atroviride infection. In this study, the structure and function of the LeDnaJ07 gene was analyzed by gene cloning and overexpression in L. edodes stress-sensitive strain YS55 via the Agrobacterium-mediated transformation method. Transformants were confirmed by qRT-PCR, fluorescence observation and Southern blotting. Overexpression of LeDnaJ07 in YS55 not only enhanced L. edodes mycelial resistance to heat stress but also facilitated mycelial growth. In the presence of heat stress, the intracellular IAA content showed a significant increase in the two LeDnaJ07 overexpression strains but only a slight change in the YS55 wild type strain. Moreover, the interaction between LeDnaJ07 and LetrpE was demonstrated via Y2H and BiFC assays. These results suggested that LeDnaJ07 may be involved in regulating IAA biosynthesis and the resistance of L. edodes to heat stresses via interacting with LetrpE.

Tricholoma matsutake is one of the edible ectomycorrhizal fungi with great economic and ecological values. Artificially mycorrhized seedlings inoculated with T. matsutake strains from Finland and Japan have been widely reported. However, few reports on the morphological characteristics of mycorrhizae synthesized from the Chinese T. matsutake strain are available to date. Here, we find that the T. matsutake strain YN1 from Yunnan province, China, can form well-developed ectomycorrhizae in symbiosis with both exotic (Pinus elliottii) and Chinese native (Pinus armandii) species. The symbiotic structures of ectomycorrhizae included obvious mantle sheaths and Hartig nets visible under a microscope. The ectomycorrhizae appeared earlier in a larger quantity in P. elliottii than in P. armandii in the culture-plate symbiotic system. The ectomycorrhizae obviously promoted the growth of potted P. armandii and P. elliottii, accompanied by the formation of shiro-like structures. Our results provide references for the effective management of ectomycorrhizae synthesis in both P. armandii and P. elliottii seedlings to form shiro-like structures, and also provide a new perspective for the afforestation and mushroom cultivation research on T. matsutake.