猴头菌属（Hericium erinaceus）菌种鉴定
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摘要
本论文从传统的生物学、生物化学和分子生物学三个方面，对猴头菌种的鉴定方法进行了比较系统的研究。国内外研究结果表明猴头菌属菌株营养丰富，含有多种活性物质，具有组织修复、增强细胞活力、延年益寿、美容健体等功能。目前，我国用于栽培生产和发酵工业的猴头菌属菌株已有的品种达几十种。它们的营养成分和药效成分各不相同，对生长环境的要求也有所差异。而且同一菌种多名和多个菌种同名的现象也不少见，这给引种、栽培和开发利用带来较大的困难。
本论文的传统生物学鉴定方法主要是利用拮抗试验，观察菌种的外部形态特征，来判定不同菌株之间亲缘关系的远近。猴头菌属之间营养菌丝的融合存在不亲合性，因此在相互作用的两个菌株的边界上，出现退化的菌丝，其背面呈浅黄色。不同品种的食用菌在生长发育中，菌丝会相互限制对方的生长蔓延，在两种不同菌落相互接触交界处出现退化菌丝，形成黄色或棕黑色的拮抗线条纹，拮抗反应不仅表现在种间，而且表现在种内品种间。根据拮抗线的宽度、颜色等特征，可以辅助进行鉴定菌种亲缘关系远近。
在生物化学方面，本论文通过对几种常用的真菌分类方法进行分析比较后，采用酯酶同工酶聚丙烯酰胺凝胶电泳，对生长在吉林省的12个猴头菌属菌株进行分析，发现各菌株在酶谱上均有丰富的表达，并呈现很强的特异性，其差异程度也各不相同。且在Rf=0.507处出现一条稳定一致的基本酶带，进一步采用模糊聚类分析法进行数据分析，表明各菌株隶属度在[0.56，0.95]之间。通过对亲缘关系树状图进行分析，得出其遗传相似性。
在分子生物学方面，本论文采用RAMP技术，对猴头菌属菌株进行DNA提取、PCR扩增及图谱分析，发现各菌株在图谱上均有丰富的表达，并呈现很强的特异性，其差异程度也各不相同。通过对亲缘关系树状图分析，得出各菌株的遗传相似性。为我省猴头菌种质保藏、知识产权保护和菌种开发利用等具有十分重要的意义。
The difference between the identification methods of the Hericium erinaceus were compared systemly in terms of traditionary biology, biochemical and molecule biology in this article. According to literature, Hericium erinaceus is rich in nutrients and contains various active substances. It can repair our tissue, enhance cell energy, lengthen life and facial and so on. Now, there are kinds of Hericium erinaceus in tragacanth and cultivate.But Hericium name is confusion identical Hericium have some names and different Hericium has identical name. It is very difficult that we plant and development and so on. Identification of Hericium with molecule biological method (RAMP) has not been reported in literature.
We can observation external morphological character and decide genetical relationship of different Hericium. Vegetative hyhae of different Hericium has disaffinity, so we can see pale yellow mycelium in interactive Hericium. At the edge of distinct Hericium have yellow or pullous degeneration hypha. But identical Hericium have not yellow or pullous degeneration hypha. Rivalry assay can show interspecies and outer species. According to width and colour of rivalry linear, we can judge genetic relationship of Hericium.
In term of biochemical, the esterase isozyme of twelve strains,which were widely cultivated in Jilin province ,were examined by plyacrylamide slab gel electrophoresis.The results show that there were plentiful expression and great specificity on isozyme patterns among strains and the degree of specificity is different,too.They had one steady identical basic isozyme band at 0.507 of Rf value.Further quantitative analysis using Fuzzy clustering indicated that the subjection degree ranged from 0.56 to 0.95 among strains.The analysis of tree figure showed there are similarity in heritability.
In term of molecular biology, we adopt RAMP in the study for indentifltion of Hericium erinaceus, which were widely cultivated in Jilin province .DNA were extracted. PCR expansion and gel. The results show that there were plentiful expression and great specificity on DNA patterns among strains and the degree of specificity is different, too. The analysis of tree figure showed there are similarity in heritability.It provides great meanings on reservation,culture and using .
本论文的传统生物学鉴定方法主要是利用拮抗试验，观察菌种的外部形态特征，来判定不同菌株之间亲缘关系的远近。猴头菌属之间营养菌丝的融合存在不亲合性，因此在相互作用的两个菌株的边界上，出现退化的菌丝，其背面呈浅黄色。不同品种的食用菌在生长发育中，菌丝会相互限制对方的生长蔓延，在两种不同菌落相互接触交界处出现退化菌丝，形成黄色或棕黑色的拮抗线条纹，拮抗反应不仅表现在种间，而且表现在种内品种间。根据拮抗线的宽度、颜色等特征，可以辅助进行鉴定菌种亲缘关系远近。
在生物化学方面，本论文通过对几种常用的真菌分类方法进行分析比较后，采用酯酶同工酶聚丙烯酰胺凝胶电泳，对生长在吉林省的12个猴头菌属菌株进行分析，发现各菌株在酶谱上均有丰富的表达，并呈现很强的特异性，其差异程度也各不相同。且在Rf=0.507处出现一条稳定一致的基本酶带，进一步采用模糊聚类分析法进行数据分析，表明各菌株隶属度在[0.56，0.95]之间。通过对亲缘关系树状图进行分析，得出其遗传相似性。
在分子生物学方面，本论文采用RAMP技术，对猴头菌属菌株进行DNA提取、PCR扩增及图谱分析，发现各菌株在图谱上均有丰富的表达，并呈现很强的特异性，其差异程度也各不相同。通过对亲缘关系树状图分析，得出各菌株的遗传相似性。为我省猴头菌种质保藏、知识产权保护和菌种开发利用等具有十分重要的意义。
The difference between the identification methods of the Hericium erinaceus were compared systemly in terms of traditionary biology, biochemical and molecule biology in this article. According to literature, Hericium erinaceus is rich in nutrients and contains various active substances. It can repair our tissue, enhance cell energy, lengthen life and facial and so on. Now, there are kinds of Hericium erinaceus in tragacanth and cultivate.But Hericium name is confusion identical Hericium have some names and different Hericium has identical name. It is very difficult that we plant and development and so on. Identification of Hericium with molecule biological method (RAMP) has not been reported in literature.
We can observation external morphological character and decide genetical relationship of different Hericium. Vegetative hyhae of different Hericium has disaffinity, so we can see pale yellow mycelium in interactive Hericium. At the edge of distinct Hericium have yellow or pullous degeneration hypha. But identical Hericium have not yellow or pullous degeneration hypha. Rivalry assay can show interspecies and outer species. According to width and colour of rivalry linear, we can judge genetic relationship of Hericium.
In term of biochemical, the esterase isozyme of twelve strains,which were widely cultivated in Jilin province ,were examined by plyacrylamide slab gel electrophoresis.The results show that there were plentiful expression and great specificity on isozyme patterns among strains and the degree of specificity is different,too.They had one steady identical basic isozyme band at 0.507 of Rf value.Further quantitative analysis using Fuzzy clustering indicated that the subjection degree ranged from 0.56 to 0.95 among strains.The analysis of tree figure showed there are similarity in heritability.
In term of molecular biology, we adopt RAMP in the study for indentifltion of Hericium erinaceus, which were widely cultivated in Jilin province .DNA were extracted. PCR expansion and gel. The results show that there were plentiful expression and great specificity on DNA patterns among strains and the degree of specificity is different, too. The analysis of tree figure showed there are similarity in heritability.It provides great meanings on reservation,culture and using .
引文
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[2] 董昌金等RAPD和酯酶同工酶技术在香菇杂交育种中的应用．食用菌学报,2000,7(1)：1-7
[3] 黄大斌等．寿宁花菇菌株的酯酶同工酶研究．中国食用菌,2000,19(3)：33-34
[4] Williams JGK,Kubelik AR,Rafaiski JA,et al.Nucleic Acides Res, 1990,18:6531-6535
[5] Mc.Dermott JM,Mc.Donald BA.Annu Rev phytopathol,1993,31:353-373
[6] John G.K.Williams,Anne R.Kubelik etc.DNA polymorphisms amplified by arbitrary primers
[7] 惠东威，陈受宜．RAPD技术及其应用．生物工程进展，1992，12(6)：1-5
[8] P.Nicholsom,H.N.Rezanoor.Use of RAPD analysis and Genetic finGerprintinG to differentiate isolates of race O,C and T of Bipolaris maydis.J.PhytopatholoGy, 1993,139:261-267
[9] MarGaret J.Jones,Larry D.Dunkle.Analysis of Cochliobolus carbonum races by PCR amplification with arbitrary and Gene-specific primers.PhytopatholoGy, 1993,83(4):366-370
[10] Zabeau M.Vos P.Selective restriction fraGment amplification.A General method for DNA finGer printinG.European patent Application 94202629.7(publication No.05348AI).Paris:European patent office. 1993
[11] Vos P.HoGers R.Blecker M et al.AFLP:a new technique for DNA finGerprintinG.Nucl.Acids Res. 1995,23(21):4407-4414
[12] 李传友等．AFLP分析中多态性扩增产物的回收、克隆及鉴定．遗传，1998,20(4)：1-4
[13] MHS.Muruke et al.Identification of mushroom mycelia using DNA technique[J].tanzanian Journal of science. 2002,28(1): 115-128.
[14] 谭琦，严培兰，詹才新．利用RAPD技术对不同地理环境下柳松姑菌株亲缘关系的分析[J]．上海农业学报．1999,15(4)：18-21．
[15] 阎培生，罗信昌，周启利．用RAPD技术对木耳属菌株进行分类鉴定研究[J]．菌物系统,2000， 19(1)：29-33．
[16] 吴学谦，李海波，魏海龙等．SCAR分子标记技术在香菇菌株鉴定上的应用研究[J]．菌物学报 2005,24(2)：259-266．
[17] Guerin-Laguette A,Matsushita N,Kikuchi K et al.Identification of aprevalent Tricholoma mastsutake ribotype in Japan by rDNA IGS1 spacer characterization . Mycol Res.2000,106 (4):435-443.
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