High-yield strain of lipopeptide, and preparation method and use thereof
A high-yielding strain, lipopeptide technology, applied in microorganism-based methods, biochemical equipment and methods, mutant preparation, etc., to achieve the effect of being conducive to separation and purification, process amplification, and good genetic stability
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Embodiment 1
[0034] Embodiment 1: Utilize normal pressure room temperature plasma mutagenesis CICC10721 to obtain new bacterial strain
[0035] 1. Activation of starting bacteria: Bacillus subtilis CICC10721 purchased from China Industrial Microorganism Culture Collection Center (CICC) was used as starting strain to streak in fresh LB solid medium, cultivated at 30°C for 18 hours, and received the order Colony into fresh LB liquid medium, culture at 30°C for 16h;
[0036] 2. ARTP mutagenesis: wash the activated cells with sterile normal saline, resuspend the cells, and properly dilute the cell concentration to 10 6 ~10 8 pcs / ml, mixed with 20% glycerol 1:1, took 20ul and spread it evenly on a sterile slide. Atmospheric pressure and room temperature plasma was used for mutagenesis. The breeding system used helium as the working gas, placed 2mm below the treatment source, set the RF power to 120W, the gas flow rate to 10slm, and the treatment time to 20s;
[0037] 3. Post-cultivation and ...
Embodiment 2
[0045] Embodiment 2: Utilize normal pressure room temperature plasma mutagenesis CICC10721 to obtain new bacterial strain
[0046] 1. Activation of starting bacteria: Bacillus subtilis CICC10721 purchased from China Industrial Microorganism Culture Collection Center (CICC) was used as starting strain to streak in fresh LB solid medium, cultivated at 37°C for 36 hours, and received the order Colony into fresh LB liquid medium, culture at 37°C for 24h;
[0047] 2. ARTP mutagenesis: wash the activated cells with sterile normal saline, resuspend the cells, and properly dilute the cell concentration to 10 6 ~10 8 pcs / ml, mixed with 20% glycerol 1:1, took 20ul and spread it evenly on a sterile slide. Atmospheric pressure and room temperature plasma was used for mutagenesis. The breeding system used helium as the working gas, placed 2mm below the treatment source, set the RF power to 140W, the gas flow rate to 16slm, and the treatment time to 60s;
[0048] 3. Post-cultivation and ...
Embodiment 3
[0050] Example 3: Fermentative production of lipopeptide surfactin by new strains
[0051] The mutant strain CCTCC M 2018747 obtained in Example 1 was inoculated into the fermentation medium according to the inoculation amount of 4% in the seed medium, and cultured at 30° C. for 72 hours.
[0052] Fermentation medium composition: 10g / L glucose, 22g / L inorganic salt, 2mg / L trace element; the inorganic salt is selected from the following mixture: phosphate, nitrate, initial pH6, sterilized at 121°C for 20min. The filling volume is 100ml / 250ml, the fermentation temperature is 30°C, the rotation speed is 160r / min, and the fermentation and cultivation time is 72h.
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