White and brown strains of Flammulina velutipes were inter-crossed. All F1 showed light-brown fruitbody, suggesting that a gene for the brown fruitbody was incompletely dominant against the white one. And backcross experiment showed that more than two genes were involved in color determination. To isolate a molecular marker linked to fruitbody color, a set of primers was designed from a sequence of clones derived by a bulked segregant analysis. These markers showed a specific band which co-segregated with brown fruitbody forming strains.
Clone Cells ; Flammulina* ; Genes, vif

Winter mushroom, Flammulina velutipes, needs low temperature during its cultivation. To save on farm costs, especially during summer, a strain adaptable to a higher or elevated-temperature must be developed. At the start of breeding program, parental strains which could endure high temperature were obtained. Seuenty four dikaryotic strains were collected and divided into four groups according to the nature of temperature. They also had different fruiting temperature. Finally we selected three brown strains ASI 4048, 4057 and 4072, and collected their spores. These selected strains can germinate even at a high temperature of 32degrees C, which were dramatically higher than the other strains. Based on these results, the new white strain adapted to mid-temperature by backcross mating was developed. Molecular markers were applied to select white fruit-body producing strains without cultivation. They showed a specific band which co-segregated with brown fruitbody forming strains in BC1F1 progenies. Selected white strains were tested under several elevated temperature conditions.
Agaricales ; Breeding* ; Flammulina* ; Fruit ; Humans ; Parents ; Spores

Whole-genome sequencing of Flammulina ononidis, a wood-rotting basidiomycete, was performed to identify genes associated with carbohydrate-active enzymes (CAZymes). A total of 12,586 gene structures with an average length of 2009 bp were predicted by the AUGUSTUS tool from a total 35,524,258 bp length of de novo genome assembly (49.76% GC). Orthologous analysis with other fungal species revealed that 7051 groups contained at least one F. ononidis gene. In addition, 11,252 (89.5%) of 12,586 genes for F. ononidis proteins had orthologs among the Dikarya, and F. ononidis contained 8 species-specific genes, of which 5 genes were paralogous. CAZyme prediction revealed 524 CAZyme genes, including 228 for glycoside hydrolases, 21 for polysaccharide lyases, 87 for glycosyltransferases, 61 for carbohydrate esterases, 87 with auxiliary activities, and 40 for carbohydrate-binding modules in the F. ononidis genome. This genome information including CAZyme repertoire will be useful to understand lignocellulolytic machinery of this white rot fungus F. ononidis.
Basidiomycota ; Esterases ; Flammulina* ; Fungi ; Genome ; Glycoside Hydrolases ; Glycosyltransferases ; Polysaccharide-Lyases

The mineral contents of the cultivation substrates, fruiting bodies of the mushrooms, and the postharvest cultivation substrates were determined in cultivated edible mushrooms Pleurotus eryngii, Flammulina velutipes, and Hypsizigus marmoreus. The major mineral elements both in the cultivation substrates and in the fruiting bodies were K, Mg, Ca, and Na. Potassium was particularly abundant ranging 10~13 g/kg in the cultivation substrates and 26~30 g/kg in the fruiting bodies. On the contrary, the calcium content in the fruiting bodies was very low despite high concentrations in the cultivation substrates, indicating Ca in the cultivation substrates is in a less bio-available form or the mushrooms do not have efficient Ca uptake channels. Among the minor mineral elements determined in this experiment, Cu, Zn, and Ni showed high percentage of transfer from the cultivation substrates to the fruiting bodies. It is noteworthy that the mineral contents in the postharvest cultivation substrates were not changed significantly which implies that the spent cultivation substrates are nutritionally intact in terms of mineral contents and thus can be recycled as mineral sources and animal feeds.
Agaricales ; Animal Feed ; Calcium ; Flammulina ; Fruit ; Pleurotus ; Potassium

Cytoprotective ability of polysaccharides isolated from different edible mushrooms was investigated on the 7-ketocholesterol-induced damaged cell line. Polysaccharide extracts from six different edible mushrooms-Flammulina velutipes, Peurotus ostreatus, Lentinus edodes, Agrocybe aegerita, Agaricus blazei, and Cordyceps militaris- were prepared by hot water extraction and alcohol precipitation. Cytoprotective ability was evaluated by measuring the viable cells of the normal embryonic liver cell line (BNL CL. 2) in the presence of 7-ketocholesterol. At 80 microgram/mL of 7-ketocholesterol, cytotoxicity was very high with a loss of 98% of viable cells after 20 h of incubation. With the addition of 200 microgram/mL of each polysaccharide isolate to the cell line containing 80 microgram/mL of 7-ketocholesterol, polysaccharide isolates from both Flammulina velutipes and Peurotus ostreatus could significantly inhibit the 7-ketochoelsterol-induced cytotoxicity in the cells. But other polysaccharide isolates were not effective in inhibiting cell damage caused by the oxLDL-induced cytotoxicity.
Agaricales* ; Agaricus ; Agrocybe ; Animals ; Cell Line* ; Cordyceps ; Flammulina ; Liver* ; Mice* ; Polysaccharides ; Shiitake Mushrooms ; Water

Flammulina velutipes was transformed efficiently by Agrobacterium-mediated transformation system. The transformation frequency was about 16% with the gill tissues of the fungal fruiting body. Southern hybridization and genetic analysis suggest that the introduced DNA was inserted onto different locations of the fungal genome, and inherited stably to the next generation via basidiospores. Transformation or gene tagging with Agrobacterium T-DNA based vector should be useful for wide ranges of genetic or molecular biological studies of the mushroom.
Agaricales* ; Agrobacterium ; Animals ; DNA ; Flammulina* ; Fruiting Bodies, Fungal ; Genome, Fungal ; Gills

Four Cladobotryum isolates were collected from four different commercially grown mushroom types infected with cobweb disease in Cheongdo-gun and Chilgok-gun of Gyeongbuk Province, Korea in 2010. The isolates were identified as C. mycophilum from Agaricus bisporus and Pleurotus eryngii, C. varium from Flammulina velutipes and Hypsizygus marmoreus. The cultural characteristics of the four isolates were investigated using potato dextrose agar (PDA) media under nine different temperatures ranging from 5~32degrees C. Rapid growth of the isolates to colony diameters of 47~82 mm was observed at conditions of 18~22degrees C. No growth was observed at 32degrees C. C. mycophilum produced a yellowish red pigment while C. varium produced a cream colored pigment after cultivation for 25 days on PDA. Phylogenetic analysis of the internal transcribed spacer region and partial 28S rDNA from the four isolates confirmed they were C. mycophilum and C. varium. Cross pathogenicity tests revealed that the two isolates of C. mycophilum were highly pathogenic toward three mushroom types, but not toward H. marmoreus. The two isolates of C. varium were less pathogenic than those of C. mycophilum, but were pathogenic toward all mushroom types evaluated.
Agar ; Agaricales ; Agaricus ; Cultural Characteristics ; DNA, Ribosomal ; Flammulina ; Glucose ; Korea ; Pleurotus ; Solanum tuberosum

This study was conducted to survey multiple factors of aversion to vegetables and preferences for vegetable-related recipes in school meal services among elementary school children in order to help develop various menus and recipes for school meal services. Questionnaire survey was carried out with the study subjects, who were 401 children in 6th grade attending elementary schools in Chonnam and Kwangju metropolitan regions. Results from the survey can be summarized as follows: 65.1% of respondents answered they try to eat vegetables and other namul side dishes served in school meal service for health. As for the frequency of taking vegetables and namul side dishes out of daily meals, 47.4% of respondents chose 'once or twice'. The reasons for aversion to vegetables in boys were taste and cooking method, while girls were taste and feeling between teeth. In boys there were no differences between regions of Kwangju and Chonnam but the tendency of aversion to vegetables was significantly high in girls. As for the aspects of vegetable aversion of subjects, 46.9% of respondents took up 'black & purple' in the unfavorable color of vegetables. 49.1% in 'bitterness' and 39.2% in 'greasiness''were in terms of the aversive taste of vegetables. The aversive vegetable recipes were 58.6% in 'raw &seasoned' and the unfavorable feeling of vegetables were 53.1% in 'squashiness'. There were differences between regions of Kwangju and Chonnam with boys in color and cooking method in girls. Results from the survey on their preferences for vegetable recipes showed that leafy vegetables like crown daisy (raw/slightly seasoned) and pak choi (broth/pot stew) fell to the most aversive category, while bean sprouts (broth/pot stew) were chosen as the most favorable one. Among root begetables bell-flowers were found to belong to the least preferred recipe, while potatoes were proven to be most preferable in terms of recipes. As for fruit vegetables and other vegetables, all respondents didn't like 'fatsia shoots' vegetable and it's cooking method and they preferred 'green pumpkins (broth/pot stew)'. In respect of mushrooms, enoki mushroom (broth/pot stew) was found most preferred and had high tendency of preferences in boys and girls in Kwangju compared with Chonnam region. The study results indicated that respondents did not show big differences in factors influencing them to be averse to vegetables and their preferences for vegetable recipes depending on regions. In order to have high preference and intake in children's diets, it needs to study in reform of menu about using namul or vegetables mixed with meats and fruits that children preferred or applying roasted and fried other less than namul.
Agaricales ; Child ; Cooking ; Crowns ; Cucurbita ; Surveys and Questionnaires ; Diet ; Flammulina ; Fruit ; Humans ; Meals ; Meat ; Solanum tuberosum ; Tooth ; Vegetables

The aim of this study was to identify and characterize new Flammulina velutipes laccases from its whole-genome sequence. Of the 15 putative laccase genes detected in the F. velutipes genome, four new laccase genes (fvLac-1, fvLac-2, fvLac3, and fvLac-4) were found to contain four complete copper-binding regions (ten histidine residues and one cysteine residue) and four cysteine residues involved in forming disulfide bridges, fvLac-1, fvLac-2, fvLac3, and fvLac-4, encoding proteins consisting of 516, 518, 515, and 533 amino acid residues, respectively. Potential N-glycosylation sites (Asn-Xaa-Ser/Thr) were identified in the cDNA sequence of fvLac-1 (Asn-454), fvLac-2 (Asn-437 and Asn-455), fvLac-3 (Asn-111 and Asn-237), and fvLac4 (Asn-402 and Asn-457). In addition, the first 19~20 amino acid residues of these proteins were predicted to comprise signal peptides. Laccase activity assays and reverse transcription polymerase chain reaction analyses clearly reveal that CuSO4 affects the induction and the transcription level of these laccase genes.
Copper Sulfate ; Cysteine ; DNA, Complementary ; Flammulina* ; Fungi* ; Genome ; Histidine ; Laccase* ; Polymerase Chain Reaction ; Protein Sorting Signals ; Reverse Transcription

We transformed the fip-fve gene into Pichia pastoris GS115 for inducible and constitutive expression to obtain feasible bioactvie recombinant Fip-fve. The fip-fve gene was cloned from Flammulina velutipes fruting body by PCR and ligated to pPIC9 to construct inducible expression vector pPIC9-FIP-fve, and promotor pgap was used to replace the paox1 to construct constitutive expression vector pPIC9-PGAP-FIP-fve. These two vectors were used to transform P. pastoris by PEG method. The fip-fve was expressed after histamine-absence screening and yeast colony PCR. The inducible expression level reached 158.2 mg/L at the fourth day and the constitutive expression level was 46.3 mg/L and 29.5 mg/L using glucose and glycerol, respectively. The SDS-PAGE and Western blotting both proved the correctness of rFip-fve, and the hemagglutination test indicats the rFip-fve's bioactivity.
Electrophoresis, Polyacrylamide Gel ; Flammulina ; chemistry ; Fungal Proteins ; biosynthesis ; Genetic Vectors ; Pichia ; metabolism ; Polymerase Chain Reaction ; Promoter Regions, Genetic ; Recombinant Proteins ; biosynthesis