Screening Of Cellulase Producing Fungus ZDTJ097-1-7from Bohai Bay And Its Yielding Conditions

Cellulase is the series of enzyme that can hydrolysis cellulose into cellobiose and Iucose,it is not a single enzyme, but the enzyme system that play Synergism.It widely exists in nature biology, such as bacteria, fungus and certain animal body. The unique marine environment contains the rich cellulose resources, this will provide important material foundation for marine microorganisms in producing Cellulase.ln view of the characteristic that marine microorganisms can produce cellulase in an extreme environment, and the fact cellulase that used in the production of the yield is still mainly comes from the fungus of land, which provide a new idea to screen the marine fungi. A filamentous fungus ZDTJ097-1-7that producing cellulase was screened out of25isolates from Bohai-Bay through Congo-red transparent circle method and cellulase activity assay. Optimum conditions were examined for the cellulase producing and catalyzing. The strain was identified based on its morphological characteristics and18SrDNA-ITS sequencing.The results are as follows:1、8strains were screened which the ratio higher than1.5through the Congo-red plate, which provides the basis for next screen.2、Collect fermented liquid after fermentation,determine the enzyme activity comprehensive results of Congo-red plate, identity the target strain.3、Optimize the conditions of fermentation and the medium components, and the results shows:When carbon source of CMC-Na concentration is4g/mL, cultured under ammonium sulfate、pH6.5、105～7Spores of fungus were incubated、150ml liqud、culture medium under150r/min for5days at24℃, under these conditions,the enzyme activity is highest.4、The study of the nature of the enzyme shows that:this enzyme perform the high activity from30℃～70℃. The filter paper cellulose plays very stable, and the activity of CMC cellulose is strongest at60℃, and the β-glucosidase has higher activity at50℃. The cellulose of ZDTJ097-1-7has the characteristics of high temperature resistance, the optimum pH is6.5for the filter paper cellulose and CMC cellulose, show neuter enzyme characteristics. And β-glucosidase plays high activity at pH8, show the alkalinity enzyme characteristics.5、Preliminary separaton arid analysis of cellulase,the result shows that:Fermented liquid precipitationed by ammonium sulfate, purified ratio improved to1.7277; Analysis the soenzyme graph of CMC cellulose.The enzyme has the heterogeneity compare with the cellulose which saled by market.The sample was detected3sozymes, the molecular mass were broader than the one saled(Fig.3-10). The enzyme activity and molecular mass size was positively relation. SDS-PAGE measured molecular mass, this enzyme system in the molecular mass range between26kDa～66.2kDa.6、The result of Classical classification show that:The fungus has the typical characteristics of the Penicillium. Homology analysis result further shows that Cellulase Producing Strain ZDTJ097-1-7from Bohai Bay was reached99%compared with the recorded penicillium though18SrDNA ITS blast in Gen-Bank database.But there exists inconsistency. Accordingly, the strain is tentatively scheduled for Penicillium ZDTJ097-1-7(Penicillium sp ZDTJ097-1-7).