Studies On Isolation, Identification, Induced Mutation Of High-Cellulase Producing Strain And Conditions Of Enzyme Production

The objective strain (a high cellulase-producing strain researched in our Labbefore) was screened from a traditional fermented food in Jiangxi province. A lot ofmonospores were isolated by the combination of plate serial dilution methods andmonospore isolation.These monospores were cultured into mature strains from whichwe got the highest cellulase-producing strain NC-1-6 by isolation many times. Weidentified this strain as Neurospora Crassa by both methods of morphological andmolecular biology identification.Effect of rice straw concentration, fermentation temperature and initial pH valueon strain growth and cellulose-producing during its growing period were estimatedwhen rice straw was used as the only carbon source. The results showed a positivecorrelation between rice straw degradation rate and strain growth status, ie 2ï¼…of ricestraw concentration was the best both for strain growth and rice straw degradation.The optimal rice straw concentration, initial pH value and fermentation temperaturewere different respectively between growths and cellulase-producing. The optimalrice straw concentration for growth and cellulase-producing were 2ï¼…and 5ï¼…respectively. The optimal initial pH value was 5.5 and 4.8 seperately. The optimaltemperature for strain growth was with not constant with a starting temperature of 25℃and a final temperature of 30℃, while the optimal temperature forcellulase-producing was fixed at 30℃.Based on the wild neurospora crassa, an induced mutant 60D7Z53 wasobtained by treatments of ultraviolet radiation and diethyl sulfate. The optimaltreating time of ultraviolet radiation or diethyl sulfate and the rate of positivemutation and lethal efficiency were assessed after each treatment. The isolation wascarried out by two steps. First isolation was according to the ratio of diameter of clearzone and that of clone (HC value); second isolation was according to cellulases'activity: filter paper enzyme (FPase) activity and CMC-enzyme (C_xase) activity afterliquid fermentation. The cellulases'activity of the mutant 60D7Z53 was much higherthan wild starting strain. They were increased by 179.25ï¼…for FPase, 175.99ï¼…forCMCase, 147.2ï¼…for C₁ase and 3.83ï¼…forβ-glucosidase respetively.The optimal culture medium and conditions of cellulase production on the mutant 60D7Z53 were carried out. The better culture medium were: 5% corn cobpowder as the carbon sources, 0.42% soybean powder as the nitrogen source, withimproved Mandels' nutrient salts, 0.1% emulsifier Tween 80 as cellulase accelerator;The better culture conditions were: 5.5 of initial pH value, 28℃of culturaltemperature, 6 days of cultural time, 3% of the quantity of inoculability, 36 h of strainage, 50 mL/250 mL of liquid volume. The optimal temperature and initial pH valuefor mutant 60D7Z53 were different and the maximal cellulases' activity appeared oneday later.