The Comparative Study Of Liquid Culture And Genetic Characteristic Of The Strain Matsutake Strains In The Different Habitat

Matsutake is a common name which contained some species of Tricholoma similar toTricholoma matsutake, it belongs to Basidiomycetes, Agaricales, Tricholomataoeae, Tricholoma,and is a kind of unique edible fungus which have highly value in the world and growsymbiotically with Pinus.and Quercus. It is mainly distributed in Asia region, parts of themappeared in America, Europe and. north of Africa. The diversities of genetic characters of matsutakewhich grows in different regions and hosts may arise by long-term systematical evolution. Currently, theresearch of concerning matsutake mainly includes general classification, identification, artificialcultivation, ecosystem management etc. There are little reports about liquid culture and geneticcharacteristic of matsutake under the different distributions and habitat condition. As a kind ofvaluable fungus, Matsutake is not cultured artificially and the wild resources are reducing, so it isnecessary to study the further genetic background analysis and the liquid culture, meanwhile, it willplay an important part in our country to promote the whole technique level of special biologicalresources, to cultivate new and special bio-industry of economical resources.To explore the appropriate factor, make clear the similarities and differences of geneticcharacteristic of matsutake in different ecological environment, select the best strains of matsutake andbest culture condition, this study applied the growth characteristic of mycelia, esterase isozyme andsmall-subunit of mitochondrial ribosomal DNA (mt-rDNA) to analyze the matsutake strains collectedfrom China, Japan, America, Sweden and north Africa which are primary distributions, growsymbiotically with different habitat. It is a basis for research and development of substituteproduction of special matsutake biological resources and produce massively. Differentdistributions and habitat genetic characteristic are different. The results are as followed:(1) The samples are divided into two groups according to its differentmorphologicalcharacters of colonies: one is Tricholoma matsutake which contains strains whichhosted with Pinus densiflora, Pinus thunbergii, Quercus sp, Pinus contorta, Tsuga sieboldii,Pinussylvestris, the codes are respectively L89-2, L89-3, 91035, Old cap, Y-N-05, 240625, 240621,Eric; The other is T.magnivelare and T.caligatum which hosted with American coniferousforest,American Pinus contorta and north African Cedrus libanotica, Am-3, Am-20 and 241803.(2) By the growth rate of mycelia and analysis of variance, we got two strains which are most suitable to grow in liquid culture, respectively are Tsuga sieboldii and PinusPinaster,240621 and Am-20.(3) Making clear the optimum condition of liquid culture: 1000ml distilled water is composed of 20gglucose, 1g typrtone and 1g yeast extract, pH 5.2～5.3, vibration culture 180 r/min of rotational speed,24^℃.(4) The suitable condition of esterase isozyme electrophorese was proposed, the concentrationgummy is 2.5% and the division gummy is 7.0%, electrophorese voltage is concentration gummy140V, division gummy 200V, electrophorese 3.5h.(5) The suitable amplification condition of mtDNA-PCR is different for the tested strains.The stains which appeared in china Pinus densiflora is: 1.5μl MgCl₂, 2μlprimers.Amplificationwas carried out as following: 30s at 94℃, 30s at 55℃. The stains which hosted with Pinusdensiflora, Pinus thunbergii, Quercus sp, Pinus contorta in Japan, coniferous forest and Pinuscontorta in America, Cedrus libanotica in the north of Africa is: 1.5μl MgCl₂, 3μl primers.Amplification was carried out as following: 50s at 94℃, 50s at 55℃, 50s at 72℃.(6) Phylogenetic tree was generated through esterase isozyme zymogram analyzed byNTSYS software. The samples are divided into three groups: one is the same as the first result ofmorphological characters of colony; the second is T.magnivelare that contained coniferous forest andPinus contorta in America; the third is T.caligatum which contains strains 241803 in Cedrus libanoticain the north of Africa.(7)The sequences of mtDNA-Ms indicate that the genetic characteristic is variable in differentdistributions and hosts: American Matsutake has obvious host diversity; Tricholoma matsutake inAsia and Europe have the high homology; the Matsutake from Africa has no obvious diversity withthe Matsutake from Asia, America and Europe.