【摘要】：多孔菌是指子實層體呈孔狀、質(zhì)地為革質(zhì)至木質(zhì)的一類大型擔子菌,主要生長在各類木材上。部分多孔菌具有產(chǎn)酶活性、藥用活性、降解活性等應用價值。開展多孔菌的采集、分離與功能活性研究,可為多孔菌資源的開發(fā)利用,奠定理論和實驗基礎。本文針對采集的野生多孔菌,開展分離鑒定、培養(yǎng)條件、抗氧化活性、胞外漆酶活性、農(nóng)藥降解作用研究。具體內(nèi)容如下:1.對兩株野生多孔菌子實體進行分離純化獲得純培養(yǎng)SS菌株和BJ菌株,并對其分類學地位、最適培養(yǎng)條件和液體發(fā)酵產(chǎn)物抗氧化活性進行分析。結(jié)果表明:結(jié)合形態(tài)學特征和ITS鑒定,確定SS菌株為多瘤木層孔菌(Phellinus tuberculosus)。菌絲體最適培養(yǎng)條件研究表明,最適碳源為葡萄糖,最適氮源為黃豆粉,最適C/N比為20/1,最適生長因子為維生素C,最適溫度為28℃,最適pH為7.0。發(fā)酵液抗氧化活性為0.102 mM(FeSO4);BJ菌株為石榴嗜藍孢孔菌(Fomitiporia punicata)。菌絲體最適培養(yǎng)碳源為葡萄糖和麥芽糖,最適氮源為酵母浸粉,最適C/N比為10/1,最適溫度為28℃,最適pH為7.0。發(fā)酵液總抗氧化活性為0.517 mmol/L(維生素E),菌體的總超氧化物歧化酶活力為770.37 U/g。2.分離獲得一株高產(chǎn)漆酶活性的菌株,編號F1635,經(jīng)過形態(tài)學和分子學鑒定為硬毛栓菌(Trametes trogii),利用DEAE-Sepharose、SP-Sepharose和凝膠過濾層析等手段,對發(fā)酵液漆酶進行純化,最終得到電泳純級的酶蛋白(TsL)。進一步開展酶學特性、降解作用、基因克隆的研究。結(jié)果表明:該漆酶是分子量為64.8 kDa的單亞基蛋白。當以ABTS為作用底物時,Tsl的最適pH為2.6,最適溫度為50℃,酶促動力學常數(shù)Km和Vmax值分別為18.58μM和1.125μmol/min。TsL漆酶對溴百里酚藍、甲基橙、鉻黑T、伊文思藍等染料具有較強的降解作用,在12 h內(nèi)降解率均達到78%以上;對農(nóng)藥乙酰甲胺磷和高效氯氰菊酯無明顯降解作用。利用簡并引物對漆酶的保守區(qū)進行克隆,得到長度分別為1246 bp和1264 bp的兩條漆酶部分基因的cDNA序列。
[Abstract]:Porous bacteria are a kind of large basidiomycetes with pore shape and leathery to wood texture, which mainly grow on all kinds of wood. Some porous bacteria have the application value of enzyme production activity, medicinal activity, degradation activity and so on. The study on the collection, isolation and functional activity of porous bacteria can lay a theoretical and experimental foundation for the exploitation and utilization of porous bacteria resources. In this paper, the isolation and identification, culture conditions, antioxidant activity, extracellular laccase activity and pesticide degradation of wild porous bacteria were studied. The details are as follows: 1. Pure cultured SS and BJ strains were obtained by isolation and purification of two wild porous bacteria fruiting bodies, and their taxonomic status, optimum culture conditions and antioxidant activity of liquid fermentation products were analyzed. The results showed that combined with morphological characteristics and ITS identification, SS strain was identified as (Phellinus tuberculosus). The optimum culture conditions of mycelium showed that the optimum carbon source was glucose, the optimum nitrogen source was soybean powder, the optimum C / N ratio was 20 渭 1, the optimum growth factor was vitamin C, the optimum temperature was 28 鈩，

本文編號：2472641