Optimization of RAPD Reaction System on Genomic DNA of Agrocybe chaxingu Huang

[Objective] The research aimed to screen out optimum RAPD reaction system on genomic DNA of Agrocybe chaxingu Huang.[Method] The single factor experiment was adopted to select the required Mg2+ concentration, template DNA concentration,primer concentration,dNTPs concentration,Taq enzyme concentration and anneal temperature initially.[Result] The optimum reaction system for RAPD amplification of Agrocybe chaxingu Huang was listed as follows:2.5 μl Buffer, 2 mmol/L Mg2+, 75 ng DNA, 0.5 μmol/L primer, 150 μmol/L dNTPs and 2.0 Taq enzyme.The reaction process was also listed as follows: denaturation for 5 min at 92 ℃,35 cycles(1 min at 92 ℃, 1 min for 35.5 ℃ and elongation for 2 min at 72 ℃),10 min at 72 ℃.[Conclusion] The research provided reference for conducting RAPD analysis of and studying genetic relationship and genetic diversity of Agrocybe chaxingu Huang.

An Orthogonal Experiment of Papain-Assisted Extraction of Polysaccharides from Agrocybe aegirit

[Objective] This study aimed to explore optimum conditions for papain-as- sisted extraction of polysaccharides from Agrocybe aegirit. [Method] Because enzyme is capable of destroying organizational structure of cells, papain was selected to cat- alyze cells to let polysaccharides out. An orthogonal experiment was carried out to investigate the effects of hydrolysis temperature, hydrolysis time, enzyme concentra- tion and solid to liquid ratio on extraction of polysaccharides from A. aegirit. [Result] Through the orthogonal experiment, the optimum papain-assisted extraction conditions were obtained： hydrolysis temperature, 45 ℃; hydrolysis time, 2 h; enzyme concen- tration 2%; solid to liquid ratio, 1：80. [Conclusion] This study will provide a reference for extraction of polysaccharides from A. aegirit.

Two novel fomannosane-type sesquiterpenoids from the culture of the basidiomycete Agrocybe salicacola

Two novel fomannosane-type sesquiterpenoids,agrocybins H(1)and I(2),together with a known compound illudosin(3),were isolated from the culture broth of the mushroom Agrocybe salicacola.Their structures were elucidated by extensive spectroscopic analysis.The relative stereochemistry of 1 was determined by the use of single crystal X-ray crystallographic diffraction.

Unusual illudin-type sesquiterpenoids from cultures of Agrocybe salicacola

Seven new illudin-type sesquiterpenoids,agrocybins A-G(1-7),along with three known analogues(8-10),have been isolated from the culture broth of the fungus Agrocybe salicacola.Their structures were elucidated on the basis of extensive spectroscopic data analysis and comparison with data reported in the literature.The relative stereoconfigurations of 1 and 6 were elucidated by the X-ray crystallographic diffraction analysis.Compound 1 was highly cyclized containing seven chiral carbons which arranged compactly in six rings.

Three New Sesquiterpenoids from Agrocybe sp.

Two new marasmone derivatives,1α,2β-dihydroxymarasmone（1） and 2β,3α-dihydroxymarasmone（2）,and one new norsesquiterpenoid,14-hydroxyoreadone（3）,were obtained from the fermentation broth of Agrocybe sp.,together with three known sesquiterpenes,agrocybolacton（4）,marasmene（5） and 14-hydroxymarasmene（6）.Their structures were elucidated by spectroscopic analyses,including 1D-and 2D-NMR experiments,high resolution electron spray ionization quadrupole time of flight（HR-ESI-Q-TOF） mass spectrometry,chemical correlation,X-ray single crystal diffraction and also compared with reported data.

Mycelium Growth and Yielding of Black Poplar Mushroom-Agrocybe aegerita （Brig.） Sing. on Different Substrates

Mycelial growth and yield of two strains ofA. aegerita on different substrates was investigated. Mycelial growth on agar media （PDA, standard, wheat, MEA, CYM, PCA, sawdust extracts： alder and mixture of beech and birch （1：1）） and sawdust substrates （birch, beech, oak, maple, alder） was investigated. Petri dishes （Ф 9 cm） for agar media and biological tubes （18 cm long and Ф 2.5 cm） for sawdust substrates were used. Yield and morphological features were studied on birch, beech and mixture of beech and alder （1：1） sawdust substrates. The temperature of inoculation for agar media and sawdust substrates was 25 ℃. For yielding, when mycelium has completely overgrown the substrate the temperature was decreased to 15-17 ℃ to initiate primordia formation. The cultivation was enlightened 10 h/d with daylight lamps （500 Ix）. One crop was harvested after five weeks. The carpophores of black poplar mushrooms were picked up in clusters. There was no statistically important difference between the mycelial growths of the investigated strains. The best growing agar media were PDA （7.3 cm）, MEA （7.2 cm） and wheat （7.1 cm）, both strains showed the slowest mycelium growth on CYM （5.2 cm）. The mycelial growth on sawdust was the best on the beech （7.1 cm） and birch （6.8 cm） sawdust. The best substrate for cultivation ofA. aegerita was mixture of beech and alder sawdust （39.5 g/100 substrate DM）. The dry yield of carpophores were the highest on beech and alder substrate （3.2 g/100 substrate DM）, but dry matter content was the highest on beech sawdust （15.7%）. The heaviest carpophores were harvested from birch sawdust （3.7 g）; the biggest caps from beech and alder sawdust （3.3 cm）. There were no statistically important differences between the mycelium growth and yielding between both investigated strains.

Fruiting Body Production and Aroma Profile Analysis of <i>Agrocybe aegerita</i>Cultivated on Different Substrates

The industrial cultivated basidiomycete Agrocybe aegerita, also known as “Pioppino” in Italy or “Samthaube” in Germany, is a high quality mushroom with a delicious aroma. Cultivation of A. aegerita on wheat straw supplemented with different residues of the food industry showed the highest yields in fruiting body production with a biological efficiency of 36% when black tea pomace was added. The addition of other substrates resulted in biological efficiencies of 23% to 33%. Besides the mushroom yields, the number and size of fruiting bodies harvested on the different substrates were determined. A comparison of the aroma profiles of A. aegerita grown on wheat straw and on wheat straw supplemented with black tea by means of GC/MS/MS/O is presented.

Adjuvant effects mediated by the carbohydrate recognition domain ofAgrocybe aegerita lectin interacting with avian influenza H9N2 viral surface glycosylated proteins

Objective: To evaluate the potential adjuvant effect of Agrocybe aegerita lectin(AAL), which was isolated from mushroom, against a virulent H_9N_2 strain in vivo and in vitro. Methods: In trial 1, 50 BALB/c male mice(8 weeks old) were divided into five groups(n=10 each group) which received a subcutaneous injection of inactivated H_9N_2(control), inactivated H_9N_2+0.2%(w/w) alum, inactivated H_9N_2+0.5 mg recombinant AAL/kg body weight(BW), inactivated H_9N_2+1.0 mg AAL/kg BW, and inactivated H_9N_2+2.5 mg AAL/kg BW, respectively, four times at 7-d intervals. In trial 2, 30 BALB/c male mice(8 weeks old) were divided into three groups(n=10 each group) which received a subcutaneous injection of inactivated H_9N_2(control), inactivated H_9N_2+2.5 mg recombinant wild-type AAL(AAL-wt)/kg BW, and inactivated H_9N_2+2.5 mg carbohydrate recognition domain(CRD) mutant AAL(AAL-mut R63H)/kg BW, respectively, four times at 7-d intervals. Seven days after the final immunization, serum samples were collected from each group for analysis. Hemagglutination assay, immunogold electron microscope, lectin blotting, and coimmunoprecipitation were used to study the interaction between AAL and H_9N_2 in vitro. Results: Ig G, Ig G1, and Ig G2 a antibody levels were significantly increased in the sera of mice co-immunized with inactivated H_9N_2 and AAL when compared to mice immunized with inactivated H_9N_2 alone. No significant increase of the Ig G antibody level was detected in the sera of the mice co-immunized with inactivated H_9N_2 and AAL-mut R63 H. Moreover, AAL-wt, but not mutant AAL-mut R63 H, adhered to the surface of H_9N_2 virus. The interaction between AAL and the H_9N_2 virus was further demonstrated to be associated with the CRD of AAL binding to the surface glycosylated proteins, hemagglutinin and neuraminidase. Conclusions: Our findings indicated that AAL could be a safe and effective adjuvant capable of boosting humoral immunity against H_9N_2 viruses in mice through its interaction with the viral surface glycosylated proteins, hemagglutinin and neuraminidase.

茶树菇多糖提取及其抗氧化性能的研究

利用水浸提法提取茶树菇(Agrocybe cylindracea)多糖,通过单因素试验考察料水质量比、浸提时间、pH、醇析时乙醇用量等4个因素对茶树菇粗多糖得率的影响。结果表明,茶树菇粗多糖的最佳提取条件为料水质量比1∶10、浸提时间2 h、pH 7.5、浸提浓缩液4倍体积的95%乙醇醇析。用乙醚和Sevage法纯化茶树菇多糖,并对其抗氧化性能进行了分析,发现茶树菇多糖具有良好的抗氧化性能。在试验浓度范围内,其总还原能力和对羟自由基、超氧阴离子自由基、脂过氧自由基的清除能力随多糖浓度的升高而增强,且其在酸性条件下对亚硝酸根离子具有良好的清除能力。

Searching Development-deficient Genes in Edible Mushroom by Self-crossing

[Objective] This study aimed to investigate the effect of mating factor recombination on the development of edible mushroom,and explore the possibility for using self-crossing to breed sporeless/low sporing mushroom strains.[Method] Selecting Agrocybe salicacola strain YAASM0711 as the self-crossing material,six self-crossed combinations were directionally constructed with single spore growth rate as an indicator,to carry out comprehensive analysis of the qualitative traits（color,deformity and growth characteristics）,quantitative traits（shape,yield,size and number of fruit bodies） and mating types of its self-crossed F1 progenies.[Result] After mating factor recombination,self-crossed single spore progenies showed serious genetic differentiation,the average mycelial growth rate and agronomic traits of inbred progenies were lower than those of the parents.The phenotype in fruiting body formation could be divided into normal fruiting and abnormal fruiting（including abnormal fruiting bodies,sporeless or low sporing fruiting bodies,no fruiting）.To be specific,18% self-crossed strains had superior agronomic traits to the parents,which can be used for selecting predominant strains from self-crossed progeny populations based on different breeding purposes.[Conclusion] The genes controlling mycelial growth rate of single spore were linked with mating factors.The mating factor-recombinant single spore strains could be selected according to the correlation between mating type and growth rate.The fruiting result of self-crossed strains indicated that the genes related to sporeless/low sporing development were recessive.Not only genetic materials harboring development-deficient genes in edible mushroom can be obtained by self-crossing,but also the sporeless/low sporing mutant strains with important application prospects can be bred.

热水法提取香菇多糖和茶树菇多糖研究

真菌多糖具有降血糖、抗肿瘤、抗氧化、抗病毒和免疫调节等多种生理功能,且对机体毒副作用小,具有广阔的开发前景。以营养价值和药用价值均较高的香菇(Lentinus edodes)、茶树菇(Agrocybe aegirit)为原料,用热水提取法在90℃、料液比1∶20(g∶m L)、提取1次、4 h的条件下,提取两种真菌多糖。结果表明,热水法提取香菇多糖的得率为9.70%,茶树菇多糖的得率为13.10%,说明茶树菇中的多糖含量高于香菇。该研究可为后续开发利用真菌多糖提供重要依据。

杨树菇栽培新技术

杨树菇（Agrocybe aegerita）又名柱状田头菇、茶树菇、茶薪菇、柳环菌、柳菇柳松茸、柱状环锈伞。属担子菌亚门,层菌纲,伞菌目,粪锈伞科,田蘑属。自然界于春秋两季多发生在杨树、柳树、榕树、小叶榕等枯干、伐桩及林间地上。其菇盖肥厚,柄脆嫩,味道鲜美,口感佳,经常食用具有抗衰老、降低胆固醇、防癌等保健功效,深受消费者青睐。

优良的栽培食用菌新品种——柱状田头菇

柱状田头菇[Agrocybe cylindracea(Dc.ex Fr)R.maire]是一种重要的野生食用菌,在欧、美、日本以及我国东北地区都十分著名,被视为野生食用菌中之佳品。该菌主要生长在阔叶树腐术上,间或亦有寄生。目前,国外尚未见栽培报道,国内曾有人做过栽培试验,但未作大面积栽培推广。

中日两种柱状田头菇的比较研究

柱状田头菇Agrocybe cylindracea,在日本称柳松茸,因常发生在柳树杆上,又具有松茸的特殊香味,故而得名。笔者曾对中国柱状田头菇作过人工驯化及栽培技术研究。1992年有幸作为共同研究员赴日本云岛县富永菌类研究所工作。在此期间作了中、日两种柱状田头菇的专题比较试验。下面介绍研究的部份内容和结果。