【作者】 刘欣；

【导师】 王秋玉；

【作者基本信息】 东北林业大学 ， 遗传学， 2009， 博士

【摘要】 活立木腐朽与木材腐朽是各国林业生产和木材使用的大敌,对腐朽前后木材成分差异和腐朽菌丝差异的研究有助于进一步理解腐朽病害发生的机理。本研究以白桦木材和5种木材腐朽菌为材料,系统分析了经5种木材腐朽菌腐朽前后白桦木材主要化学成分的变化,5种主要木材腐朽菌在生长和酶学特性的差异,以及分离了2种木材腐朽菌腐朽前后菌丝差异表达的基因,探讨了5种木材腐朽菌的生物学特性与其腐朽白桦木材能力之间的关系,从腐朽前后木材腐朽菌和白桦木材化学成分两方面的变化规律,研究二者之间的相互作用机理,并在此基础上探索了环境友好的耐腐和易腐白桦育种策略---木腐菌的腐朽能力和白桦木材的天然耐腐性基础上的选择育种。主要研究结果如下：(1)用白囊耙齿菌(Irpex lacteus)、黄伞(Pholiota adiposa)、彩绒革盖菌(Coriolus versicolor)、木蹄层孔菌(Fomes fomentarius)和桦剥管菌(Piptoporus betulinus)5种木材腐朽菌,分别对帽儿山林场的300株白桦木材进行木材天然耐腐性检测,结果表明白桦木材重量损失率株间差异极显著,在群体水平上符合正态分布。彩绒革盖菌腐朽白桦木材的能力最强,其次是木蹄层孔菌和桦剥管菌,而白囊耙齿菌和黄伞腐朽白桦木材的能力最弱。黄伞和木蹄层孔菌腐朽后的木材重量损失率呈极显著的正相关,白囊耙齿菌和桦剥管菌腐朽后的木材重量损失率呈极显著的负相关。(2)比较了腐朽木材与新鲜木材在主要化学成分上的差异,不同木材腐朽菌对木材中各主要成分的腐朽程度都不相同,说明木材腐朽菌对木材各成分有偏好性。根据白桦木材腐朽后的重量损失率,每种木材腐朽菌筛选出10株耐腐和易腐白桦,分别组成五组耐腐和易腐群体,对应群体间的木材重量损失率差异极显著。5种耐腐群体可以用于今后白桦建筑材和单板材的良种选育。发现不同木材腐朽菌筛选出来的耐腐和易腐白桦植株绝大部分都不相同。(3)分别5种木材腐朽菌将白桦木材的天然耐腐朽程度分为耐腐、易腐和中等腐朽三个层次,为今后判断白桦木材是否抗这5种木材腐朽菌腐朽提供了便利的比较方法和比较标准。(4)对筛选出的白桦耐腐和易腐植株的新鲜木样主要化学成分进行Pearson相关性分析,发现白桦耐腐和易腐群体木材中的1%NaOH抽出物和水分与纤维素含量成显著负相关,易腐群体白桦木材中的水分与纤维素含量、苯醇抽出物和总酚含量、木质素和总酚和总黄酮含量成显著正相关,这些相关关系与对应成分的特性以及在耐腐和易腐群体中的含量相符。(5)用分光光度法检测了白桦木材中的酚类和黄酮类物质含量,发现总酚和总黄酮含量显著性高的白桦木材抗木蹄层孔菌,总酚含量显著性高的白桦木材抗桦剥管菌。用GC-MS法进一步检测白桦木材,得到11种化合物。其中4,7-亚甲基-1H-茚-6-酚、N,N’-二亚水杨基-1,2-二氨基乙烷、3,4,5-三甲氧基苯酚属于酚类物质,反式-3-甲基-4-甲硫基-查耳酮属于黄酮类物质。(6)系统地比较5种木材腐朽菌在固体、液体培养基中的生长特性、木质素降解酶活性与它们降解白桦木材的能力,发现5种木材腐朽菌对白桦木材的腐朽能力与它们在液体培养基中能达到最快生长速度所需要的时间相符,与其生长速度和木质素降解酶活性都无明显相关性,即生长迅速的木腐菌并不代表其木材降解能力强。(7)木蹄层孔菌腐朽后的白桦木材1％NaOH抽出物含量最低,苯醇抽出物含量中等,纤维素含量最高,木质素含量比较低,而且其腐朽白桦木材的能力较强,适宜改造成造纸业原料处理的工程菌。彩绒革盖菌的腐朽白桦木材能力最强,但腐朽白桦木材后的纤维素含量远低于其它3种菌,因此不适于生物辅助造纸,但其锰过氧化物酶和漆酶活性最高,可以用于其它污染物治理。木蹄层孔菌筛选出的白桦易腐群体新鲜木材的1％NaOH抽出物含量中等,苯醇抽出物含量最低,纤维素含量比较高,木质素含量最低,因此最适用于碱法制浆过程的造纸。(8)黄伞同时表达木质素过氧化物酶、锰过氧化物酶和漆酶的活性,腐朽后木材的木质素损失率高于纤维素损失率,腐朽木材近似于白色,不易粉碎。黄伞应该属于白腐菌。(9)分别对白囊耙齿菌和桦剥管菌进行木材诱导,找到白囊耙齿菌的漆酶等5个同源片段和桦剥管菌的mRNA前体拼接因子syf2等7个同源片段。这些片段均在木材诱导真菌中有不同程度的表达,并且在木材腐朽菌腐朽木材过程中起了重要作用。发现白腐菌和褐腐菌在白桦木材诱导下存在明显差异表达的基因,这些差异表达片段对木材腐朽机理的研究起重要作用。更多还原

【Abstract】 Live stumpage decaying are disadvantages in international forestry production and wood utilization. The study on change of wood component before and after wood rotted and gene different expression of the rot fungi would help to understand the decay mechanism of the wood. In this study, the wood of white birch and 5 wood rot fungi were used as materials, the changes in main chemical components of the wood before and after the wood decayed by rot fungi was systematically analyzed. Different expressed genes of wood rot fungi induced by wood chip were isolated. The relationship between the biological characteristics of several wood rot fungi and their degradation ability to birch wood were discussed. The interaction mechanism between wood rot fungi and major components of the birch wood before and after decayed were investigated according to their change pattern. A environment friendly breeding strategy of the birch with susceptible and resistant characteristics was investigated, which is a selective breeding on the basis of the degradation ability of the fungi and the natural decay resistance of white birch. The major results are as follows:(1) Irpex lacteus, Pholiota adipose, Coriolus versicolor, Fomes fomenta-rius and Piptoporus betulinus were separately used to decay 300 wood samples of white birch from Maoer Shan area for testing natural decay resistance f the white birch. The differences of weight loss of decayed wood between individual trees are statistically significant. They show normal distributions at the population level. The decomposing power of Coriolus versicolor to wood samples is higher than that of Fomesfom entarius and Piptoporus betulinus, while the decomposing power of Irpex lacteus and Pholiota adipose are the weakest ones. The weight loss of decayed wood between Fomes fomentarius and Pholiota adiposa shows significant positive correlation. But the weight loss of decayed wood between Irpex lacteus and Piptoporus betulinus shows highly significant negative correlation.(2) The differences between rotten wood and fresh wood in terms of the main chemical components are compared. Different wood rot fungi has different decaying value in main chemical components of the wood, that means that different wood rot fungi have their own preference on each chemical component of wood.10 susceptible plants and 10 resistant plants were selected by each wood rot fungi based on weight loss rate of decayed wood, that grouped 5 susceptible populations and tolerance populations separately. The difference of decayed wood weight loss was the most significant between corresponding groups.5 tolerance populations could breed for construction wood and plywood. Susceptible population could breed for pulpwood. Susceptible and tolerance birch plants selected by different wood rot fungi mostly were found to be not the same. (3) To divide natural resistance ability of white birch to three levels, such as resistant, susceptible and the moderate by 5 wood rot fungi separately. It provided a convenient comparison method and criterion to judge whether the wood of a birch plant whether or not to resists one of the 5 wood rot fungi in future.(4) After the pearson correlation analysis of major wood chemical components between tolerance and susceptible populations, it was found that the 1% NaOH extraction in wood had negative correlations with the water content and cellulose. The water content and cellulose, benzene-alcohol extraction and total phenol, total phenol and total flavonoids as well as lignin have positive correlations in the wood of susceptible populations. These correlations was consistent with the related characteristics of the corresponding components, as well as with their contents in the both populations.(5) Phenols and flavonoids content in birch wood were tested by spectrophotometry. The wood with higher total phenols and total flavonoids content was hard to be decayed by Fomes fomentarius, and the wood with higher total phenols content was more inappropriate by Piptoporus betulinus. The birch wood powders were tested by GC-MS.11 compounds were discovered. 4,7-methano-1H-indenol-hexahydro,2,2’-[1,2-ethanediylbis (nitrilomethylidyne)] bis-phenol and 3,4,5-trimethoxy-phenol belong to phenols, and trans-3’-Methyl-4-(methylthio)chalcone belong to flavonoids in birch wood.(6) The growth characteristics of 5 wood rot fungi in solid and liquid media were systematically compared with the activities of ligninolytic enzymes and wood-decaying capacities. The wood-decaying capacities of white birch by 5 wood rot fungi were corresponded with the time needed to achieve the fastest growth rate of the fungi in liquid medium; but no obvious correlation with their growth rate or the activities of ligninolytic enzymes. The wood rot fungus with fast growth rate did not really show strong wood-decaying capacity.(7) For the wood decayed by Fomes fomentarius,1%NaOH extraction content was the lowest, and benzene-alcohol extraction was in the middle, and cellulose was the highest, and lignin was lower, but, which has stronger wood-decaying capacity. It means that Fomes fomentarius could be transformed into the best engineering strain for processing raw materials in paper industry.The wood-decaying capacity of Coriolus versicolor was the strongest among all 5 fungi, but the cellulose content in wood decayed by it was lower. It means that Coriolus versicolor was not suitable for bio-assisted paper-making. Meanwhile, Coriolus versicolor could be used for pollutant control since the activities of its MnP and Lac were the highest.The fresh wood of susceptible populations selected by Fomes fomentarius, which 1% NaOH extraction content was in the middle, benzene-alcohol extraction was the lowest, cellulose was highest, and lignin was also the lowest, could be the best raw materials in alkaline pulping process by using alkaline method.(8) The activities of Lip, MnP, Lac could be tested in Pholiota adiposa, which lignin loss rate was higher than the cellulose loss rate in wood decayed by it. At the same time, color of the wood decayed by Pholiota adipose looks white and the decayed wood was difficult to be crushed. Pholiota adiposa should belong to white rot fungus.(9) White rot fungus—Irpex lacteus and brown rot fungus—Piptoporus betulinus were induced by wood powder separately.5 homologous fragments, such as Laccase, etc. were found in Irpex lacteus cDNA fragments,7 homologous fragments, such as pre-mRNA splicing factor syf2, etc. were found in Piptoporus betulinus cDNA fragments. Those fragments all expressed in fungus in different level and devoted for wood decaying.The gene expressed obviously different between white rot fungi and brown rot fungi induced by wood powder of white birch. These differential expression gene fragments would play important roles in the research of wood decay mechanism.更多还原