白腐菌紫外诱变选育高产漆酶突变株及其产酶条件研究

以白腐菌为出发菌株,利用紫外线(UV)进行诱变,筛选高产漆酶突变菌株。通过测定致死率绘制出发菌株的致死曲线,采用PDA-RBBR平板变色法进行初筛,ABTS检测酶活对突变株进行摇瓶复筛。结果表明:利用15 w紫外灯在照射距离为30 cm,照射时间为120 s,致死率为72.1%的条件下进行诱变处理,获得一株高产菌株,其酶活提高79.54%,经过5代传代培养,未见酶活下降,具有较好的遗传稳定性,进一步研究了初始pH值,接种量和培养基装液量等对诱变菌株产酶的影响,结果表明在最佳的培养条件pH值6.0,15%的装液量于28℃下,酶活达214.9 U/L。     

通过碳源与氮源研究白腐菌产漆酶和菌丝体生长的相关性

通过碳源与氮源对白腐菌产漆酶和菌丝体生长的影响 ,定性地研究了白腐菌产漆酶和菌丝体生长的关系。碳源中淀粉最能提高菌丝体的生长量 ,麦芽糖是促进漆酶分泌的最好碳源。氮源中的酒石酸铵能较好地促进漆酶的分泌 ,但几种氮源对菌丝体生长的影响不是很明显。研究显示白腐菌的生长和酶的分泌不同步 ,也不成正相关。     

白腐菌产漆酶的纯化及部分酶学性质

对白腐菌W 1产生的漆酶粗酶液通过超滤浓缩、分子筛和离子交换层析进行纯化 .用SDS PAGE证明该酶的分子量大约为 6 2 4kD .等电聚焦电泳显示该酶的等电点为 3 5 .酶反应的最适温度为 5 0℃ ,最适pH值为 4 5 .此酶氧化DMP的Km 值为 3 84× 10 -5mol L .金属离子对酶活的影响很大 ,其中K+ 、Mn2 + 、Ag+ 对酶活有促进作用 ,Fe2 + 、Fe3 + 、Hg2 + 、Co2 + 、Ba2 + 等对酶活有明显的抑制作用 .酶对部分染料也有一定的脱色效果     

自腐菌漆酶的研究进展

自然界中术素的降解主要是通过丝状真菌,其中主要由白腐担子菌的分解作用来完成。白腐菌降解木质素,是通过其分泌的酶的作用来实现,白腐菌所分泌的木素降解酶主要有三种,即本素过氧化物酶（LigninPeroxidase,简称LIP）,依赖锰的过氧化物酶办hganes于depeMentoroxM用已简称MnP和漆酶（Ulccase。漆酶是一种含铜的多酚氧化酶中Aiphenoloxidase,ECI10．3．2）,最早是从漆树的分泌物中发现（Yoshi他1883）,随后人们发现一些高等真菌也能分泌这种酶（sertranct,lsoe;totrircte,is00）。现在人们知道,漆酶广泛地存在于担子菌、…     

毛栓菌(Tramets trogii)液体培养产漆酶的研究

实验研究了碳源、氮源和综合因素对毛栓菌(Tramets trogii)液体培养产漆酶和生长情况的影响。碳源为玉米面、氮源为豆饼粉时,菌株的产漆酶酶活最高;碳源为可溶性淀粉,氮源为麦麸时,对菌株的生长有利;在液体条件下,菌株产漆酶的最佳培养基组成为玉米面3.5%,豆饼粉4.0%,KH2PO40.3%,MgSO40.15%,VB10.04%。     

白腐菌在两种固体基质上的漆酶产量及同工酶活性比较研究

一株新分离的白腐菌Z1可在两种农业废弃物(麸皮和稻草秸杆)基质上生长并分泌漆酶,其中在麸皮基质上的漆酶产量比在稻草秸杆基质上的漆酶产量高出10倍左右,同时活性电泳显示Z1菌在麸皮基质上可产生3条漆酶同工酶,比在稻草秸杆基质上多1条,其活性也比在稻草秸杆基质中的漆酶活性高。     

层孔菌产漆酶的摇瓶最适培养条件研究

筛选到一株高产漆酶的层孔菌W－1,对其兴体产漆酶的培养条件进行了优化,确定了最适的碳源、氮源,并用正交试验确定了培养基中碳源和氮源的浓度。在最优化的培养条件下,培养7d后,W－1产生的漆酶活力可以达到7．1U／mL。采用补料的方法,可以得到大量的漆酶粗酶液。     

白腐真菌漆酶的纯化及性质

液体发酵培养白腐真菌F9,粗酶液经盐析、透析浓缩、葡聚糖G-100柱层析、DEAE-纤维素离子交换层析四步分离纯化,得电泳纯漆酶。经SDS-PAGE法测定酶的相对分子质量约为6×104,酶活回收率达46.47%,纯度提高了18.86倍。F9漆酶最适反应温度为40℃,最适反应pH为4.8,在35℃以下、pH 4.8～5.4的范围内稳定性较强。其催化愈创木酚的Km为4.61 mmol/L,vm为6.27 mmol/(L.min)。K+对其有激活作用,而Fe2+、Fe3+对其有明显抑制作用。     

白腐菌产锰过氧化物酶条件的研究

白腐菌ＰｈａｎｅｒｏｃｈａｅｔｅｃｈｒｙｓｏｓｐｏｒｉｕｍＭＩＧ．３８３降解桉木时具有显著的选择性,３０天内降解３７．２３％Ｋｌａｓｏｎ木素,７．２９％综纤维素。该菌株产胞外锰过氧化物酶,并在高碳低氧培养基中显示较高酶活,静置液体培养的优化培养条件是（Ｌ＾－１）：１０ｇ葡萄糖,２ｍｍｏｌ酒石酸铵,１０ｍｍｏｌｐＨ４．５醋酸钠缓冲液;１ｇ吐温８０,２ｇＫＨ２ＰＯ４,０．５ｇＭｇＳＯ４．７Ｈ２Ｏ,０．     

不同木质纤维素基质上白腐菌降解特性的研究

通过测定木质素、纤维素、半纤维素和漆酶分泌的变化,研究白腐菌在稻草、木屑、粗纤维素、滤纸、黑液木素基质上的降解特性。结果表明,除黑液木素上白腐菌不能生长外,在前25d,各基质中纤维素、半纤维素和木质素含量呈持续下降趋势,之后,降解速率减少,其中木质素的降解速率大于纤维素和半纤维素的降解速率。漆酶分泌在生长初期呈快速上升趋势,第10d酶活达到最大,第10～20d快速下降,其后基本不变,基质中酶活大小顺序为稻草基质、木屑基质、粗纤维和滤纸基质,显示了木质素存在对漆酶分泌的诱导作用。     

1株产漆酶白腐真菌的筛选和鉴定

从不同的生境采集生物样品,利用Bavendamm反应反复筛选产漆酶的白腐真菌。利用真菌通用引物对ITS1/ITS4扩增菌株rDNAITS区序列,对扩增产物进行测序。测序结果在GenBank中进行同源性搜索,下载部分具有代表性种的ITS序列进行序列比对,利用软件MEGA4构建分子系统发育树,通过序列分析,并结合形态学鉴定出4220为香栓孔菌(Trametes suaveolens)。     

白腐菌选择性降解竹基质中木质纤维素的研究

对竹基质白腐菌选择性降解进行了初步研究。结果表明,菌株B1对竹基质中木质素和半纤维素有明显的降解选择性。降解55 d木质素和半纤维素降解率分别达44.4%和47.1%;降解20 d降解选择性最好,木质素和半纤维素降解率选择系数分别是2.08和1.98。从FTIR图谱中木质纤维素相关谱峰(2 924、1 6351、6011、5101、165、1 045、666/cm等)的明显变化也可以得出相同结论。     

Species-specific Cd-stress Response in the White Rot Basidiomycetes Abortiporus biennis and Cerrena unicolor

The effect of cadmium (Cd) on fungal growth, Cd bioaccumulation and biosorption, and on the formation of potential heavy metal response indicators such as thiols, oxalate, and laccase was investigated in the white rot fungi Cerrena unicolor andAbortiporus biennis. Only the highest Cd concentration employed (200 μM) inhibited growth of C. unicolor, whereas already lower Cd concentrations caused decreasing mycelia dry weights in A. biennis. Cd biosorption onto the mycelial surface was the predominant Cd sequestration mechanism in C. unicolor. Surface-bound and bioaccumulated Cd concentrations were essentially in the same range in A. biennis, leading to considerably higher intracellular Cd concentrations in A. biennis than in C. unicolor. Oxalate and laccase were produced by both of the fungal strains and their extracellular levels were elevated upon Cd exposure. Oxalate concentrations and laccase titres were considerably higher in C. unicolor than in A. biennis. Both fungi responded to increasing Cd concentrations by increasing intracellular amounts of thiol compounds (cysteine, γ-glutamylcysteine, glutathione in both its reduced and oxidized form) but Cd application increased the amounts of thiols to a higher extend in A. biennis. Taken together, these species-specific responses towards Cd suggest that C. unicolor possesses a more efficient system than A. biennis to keep intracellular Cd concentrations low.     

Laccase isoforms with unusual properties from the basidiomycete Steccherinum ochraceum strain 1833

Aims: To isolate and characterize the laccase isoforms from S. ochraceum 1833 – a new active producer of high extracellular laccase activity. Methods and Results: Three laccase isoforms (laccases I, II and III) with 57·5, 59·5 and 63 kDa molecular masses respectively were purified from S. ochraceum 1833 and in contrast to the known laccases had strongly pronounced absorption at 611 nm with molar extinction coefficients ranging from 7170 to 7830 mol^?1 l cm^?1. All isoforms showed maximal activity with ABTS at low pH (≤2) and temperatures in the range 70–80°C, were stable for long time of incubation at high temperature (60–80°C) and at pH values ranging from 2 to 6. Laccase II showed a higher activity and wider substrate specificity. N‐terminal amino acid sequence analysis of the purified laccase II (VQIGPVTDLH) showed 80% identity with the N‐terminal amino acid sequence of laccase from Lentinula edodes [Appl Microbiol Biotechnol 60 (2002) 327]. Conclusions: Elevated temperature optima, high thermo‐ and pH‐stabilities, the broad substrate specificity of the isoforms make the laccases from S. ochraceum 1833 a suitable model for biotechnological processes proceeding at high temperatures. Significance and Impact of the Study: For the first time, new basidiomycete strain S. ochraceum was reported as a producer of novel thermostable, pH stable, acidophilic laccases with unusual spectral properties.     

紫外、离子注入法对白腐真菌诱变效应的初步研究——多酚氧化酶菌株的选育

对白腐真菌F4孢子悬液进行紫外、N+离子注入诱变.诱变后待孢子长出单菌落,滴加茴香胺等多酚氧化酶底物,观察其颜色变化;经发酵筛选,获得一株多酚氧化酶高产菌POP5,漆酶活力是原出发菌株的16倍,并且得到一株多酚氧化酶缺失菌株POL1.紫外诱变,孢子浓度为106～108个/ml,照射时间1～2min;N+离子注入,孢子浓度为105～106个/ml,能量20Kev,剂量为5×1014ons/cm2,每个平板上生长30个左右菌落是最佳诱变选育条件.与其它真菌的孢子相比,N+离子注入法对白腐真菌F4孢子的致死率较大.