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Isolation of Silkworm Sericin and Feather Degragation Bacteria and Optimization of Keratinase Fermentation Medium

Author: ZhangShaSha
Tutor: LiuXunLi
School: Shandong Agricultural University
Course: Microbiology
Keywords: The sericin and feathers keratin Degrading bacteria Separate Keratinase Ferment
CLC: X172
Type: Master's thesis
Year: 2010
Downloads: 80
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Abstract


Yishui County, Linyi City, Shandong Province, Huayin silk factory and Shandong Province, Taian Thai silver factory wastewater samples using selective medium bacteria of Silk Sericin keratin degradation Isolation and Screening. Repeated bacterial genome sequences of the PCR technology (REP-PCR), a cluster analysis of the bacterial degradation sericin capacity. Colony morphological characteristics, physiological and biochemical characteristics and 16S rRNA sequence homology analysis identified both degradation Silk sericin can be bacterial degradation of feather keratin 10. The organic solvent treatment keratin substrate was measured the the feather keratin degrading bacteria keratinase activity, and select the highest keratinase activity of Bacillus (Bacillus pumilus) S59 production keratinase optimization of the fermentation medium. The main results are as follows. Wastewater pool separated from the system Sichang 39 bacterial degradation of silk sericin isolates using Coomassie Brilliant Blue G-250 Determination of the degradation activity of the isolates sericin, the degradation rate of between 2.69 % to 28.86%. REP-PCR results of cluster analysis of the isolates showed that the dissimilarity in REP-PCR Spectrum 0.9 percent level for clustering 12 groups. 39 silk glue protein degradation bacteria isolated 10 feather keratin degrading bacteria. 10-degrading bacteria keratinase activity between 7.06 U ~ 13.49 U. Characterized by colony morphology, physiological and biochemical characteristics and 16S rRNA base sequence determination and homology analysis identified 10 bacteria are of the genus Bacillus (Bacillus sp.). 16S rRNA sequence of the strains have been registered in GenBank. Through a single single-factor test to determine the best the S59 strains producing keratinase shake flask fermentation medium carbon source, nitrogen source, carbon and nitrogen ratio, and inorganic salts. Medium optimization process, on the basis of the univariate analysis, the first fractional factorial test analysis of medium components corn flour, glucose, feathers and CaCO 3 keratinase activity, the results show that corn flour and feathers as the main influence factor, and the center point of the level of enzyme activity are negative effect, therefore, to improve the enzymatic activity required to reduce their dosage; glucose and CaCO 3 on the enzyme activity not significant, the low level of dosage as the optimal level of enzyme production. Then using a central composite design and response surface analysis to determine the optimal concentration of the main influencing factors. S59 strain producing keratinase fermentation medium: glucose 5.00 g, 32.83 g, corn flour feathers 10.67 g the CaCO 3 3.00 g, distilled water 1000 mL. Flask fermentation using the optimal fermentation medium, the fermentation broth angle protease activity reached 41.04 U, the enzymatic activity of the fermentation broth of the more basic fermentation medium (13.49 U) increased by 204.23%.

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