Method for producing high-activity chitosanase preparation
A technology of chitosanase and high activity, which is applied in the field of microbial fermentation technology and enzyme engineering, can solve the problems of long enzymatic hydrolysis time, low concentration of enzymatic hydrolysis substrate, low enzyme activity, etc., and achieve the improvement of enzyme activity and enzymolysis The effect of high process efficiency
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Embodiment 1
[0026] Example 1 Breeding of TQK strains highly expressing chitosanase.
[0027] Based on the TQS6 strain obtained from CCTCC (preserved by the China Center for Type Culture Collection), through UV (ultraviolet mutagenesis) and NTG (nitrosoguanidine mutagenesis) compound mutagenesis, the conditions for ultraviolet mutagenesis are: UV lamp power 15 ~30w, irradiation distance 20~50cm, irradiation time 1~8min. The conditions for NTG mutagenesis are: NTG concentration 1-3 mg / ml, in tris buffer solution with pH 6.0, treatment for 90-120 min. The strains were inoculated on plate medium containing 0.3% Congo red, induced and cultured at 35°C for 30 hours, and transparent hydrolysis circles could be seen. The medium formula is calculated by mass percentage: 2% agar, 0.5% chitosan, 1% tryptone, 1% yeast extract powder, 0.5% NaCl, and 0.3% Congo red.
[0028] Select the strain with the larger H / C colony diameter ratio of the hydrolysis circle diameter, carry out shake flask re-screeni...
Embodiment 2
[0029] Embodiment 2 fermentation produces chitosanase
[0030] 1): The TQK strain (preservation number is CCTCC No: M207129) was inoculated by streaking on a flat surface containing 1% yeast extract powder, 1% tryptone, 0.5% sodium chloride, 0.5% chitosan, and 2% agar Incubate at 33°C for 24 hours to use.
[0031] 2) First-level liquid culture: use 1000ml triangular flask to pack 200ml liquid medium containing 1% yeast extract powder, 1% tryptone, 0.5% sodium chloride, 0.5% chitosan, sterilize for 20 minutes at 10 pounds, and inoculate one In a ring, culture on a shaker (260r / min) at 34°C for 27 hours.
[0032] 3) Use a 7-liter fermenter to pack 5 liters of liquid medium containing yeast extract powder, tryptone, chitosan, inorganic salt composite components, and corn dregs. After the same sterilization as above, connect 200 mL of shaker flask seeds and ventilate and stir at 35 ° C. 18-24 hours out of the tank.
[0033] 4) Low-temperature centrifugal separation of enzyme li...
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