Isolation And Identification Of Cellulose-degradation Bacteria From Panda And Cow Dung

This experiment take advantage of panda’s excrement and cow’s dung to screen cellulose-degradation bacteria for exploring the panda’s digestive tract flora structure and the interaction between different species.Morphological observation, strains of physiological and biochemical test, the strain of molecular identification are used to find efficient cellulose-degrading bacteria and lay the foundation for high fiber feed utilization.The first experiment, the screening of cellulose-degradation bacteria:The screening of cellulose-degradation bacteria include two steps.Initial screening process is to put the samples into the liquid fermentation medium for training 3d by constant temperature and shocking, then fermented liquid will be diluted and coating in the solid medium plate,put the strain into a clean environment without other strain and coat the strain into the solid medium plate.choosing the larger circle of the transparent hydrolysis strains to save for subsequent research by the Identification of a red dye.the second screening is mainly to test the CMCase and FPAase’s enzyme activity and select the strain of high enzyme activity for subsequent identification.This experiment respectively screen 5 samples of cellulose degradation bacteria strains from the panda feces samples and cow dung.The second experiment, the identification of cellulose-degrading bacteriaIdentification of cellulose-degradation bacteria including three aspects,morphology observatio n,physiological and biochemical test and molecular biology identification.Morphological observati on mainly includes a gram stain and observation.the shape of bacteria, the color of colony, shape, and the size of the colony, transparency, gloss, texture, edge character, uplift shape, etcPhysiologi cal and biochemical test with reference to berger’s bacteria identification manual relevant methods of physiological and biochemical determination,including the sugar alcohol fermentation experime nts, starch hydrolysis experiments, V-P experiment, methyl red and oxidase test,catalase test (the contact enzyme test), nitrate reduction reaction, etc.Molecular biology identification mainly comp are the gene sequence and Genbank pool toidentify the kinds of strains by polymerase chain reacti on.The five dentified strains are streptomyces albus(NC020990), pseudomonas (NC020209),bacill us (NC021171), klebsiella bacteria (NC016612), paenibacillus (NC012914)from panda faeces and the other strains are pseudomonas (NC020209), volunteers hayes bacteria (NC007384), pineapple bacteria (NC013956), eosinophilic malt oligotrophic (NC010943), paenibacillus(NC012914).The third experiment, optimize the determination of the enzyme activity.The influence of the factors of enzyme activity are,cultivation time, temperature, pH, inoculation fluid volume.which are used to optimize the experimental conditionsand the activity of of the stra in through single factor.The panda’s excrement in the screening of strains for producing enzyme and enzyme production results shows that:streptomyces albus (NC020990) reach the maxium enzy me activity that is 24.732 U with the pH of 7, temperature of 35℃, liquid loading quantity of 30 %, the quantity of 2.5%, table speed is 180 RPM,culture time of 36h pseudomonas (NC020209) reach the maxium enzyme activity that is24.362 U with the pH of 6, temperature of 25℃, liquid loading quantity of 30%, the quantity of 4%, table speed is 180 rpm,culture time of 36h.bacillus (NC021171)reach the maxium enzyme activity that is23.125 U with the pH of 6.5,temperature of of 30℃, liquid loading quantity of 30%, the quantity of 2%, table speed is 180rpm,culture time of of 30h.klebsiella bacteria (NC016612)reach the maxium enzyme activity that is 18.642 U with the pH of 6.5, temperature of 30℃, liquid loading quantity of 30%, the quantity of 2%, table speed is 180 rpm.culture time of 42h. paenibacillus (NC012914)reach the maxium enzyme activity that is 9.234 U with the pH of 7, temperature of 30℃, liquid loading quantity of 30%, the quantit y of 2%, table speed is 180 rpm.culture time of 48h.The cow’s excrement in the screening of strains for producing enzyme and enzyme productio results shows that:pseudomonas (NC020209)reach the maxium enzyme activity that is23.156 U with the pH of 6, temperature of 25℃, liquid loading quantity of 30%, the quantity of 4%, table sp eed is 180 rpm,culture time of 36h.Volunteers hayes bacteria (NC007384)reach the maxium enzy me activity that is21.842 U with the pH of 7, temperature of 35℃, liquid loading quant ity of 40%, the quantity of 2.5%, table speed is 180 rpm,culture time of 42h.pineapple bacteria(NC013956) reach the maxium enzyme activity that is10.624 U with the pH of 6.5, temperature of 30℃.liquid 1 oading quantity of 30%, the quantity of3%, table speed is 180 rpm,culture time of 36h.osinophilic malt oligotrophic (NCO 10943)reach the maxium enzyme activity that is 9.942 U with the pH of 6. 5, temperature of 35℃, liquid loading quantity of 30%, the quantity of 2.5%, table speed is 180 rp m,culture time of 36h.paenibacillus(NC012914)reach the maxium enzyme activity that is 9.146 U with the pH of 7, temperature of 30℃, liquid loading quantity of 30%, the quantity of 2%, table sp eed is 180 rpm,culture time of 48h.