Method for preparing high temperature-resistant lipase

A technology of lipase and high temperature resistance, applied in the field of fermentation, can solve the problems of complex process, low enzyme yield, poor temperature resistance of lipase, etc., and achieve the effect of wide pH range, expanded application field, and good tolerance

Active Publication Date: 2016-07-27
HUNAN NANBEIWANG BIOTECH CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Plants such as the seeds of oil crops, animals such as the pancreas and adipose tissue of higher animals are the source of lipase extraction method, the extraction method has limited resources, complicated process, and low yield; microbial fermentation method is not affected by the environment, resources Abundant, short enzyme production cycle, relatively simple product and low cost, easy to manage in production, easy to realize industrial production, but the temperature resistance of lipase prepared by microbial fermentation method is relatively poor
[0004] Bacillus is an important member of li

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  • Method for preparing high temperature-resistant lipase
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  • Method for preparing high temperature-resistant lipase

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[0036] Example 1:

[0037] A preparation method of high temperature resistant lipase of the present invention, comprises the following steps:

[0038] (1) Gene cloning process: the high temperature resistant lipase gene derived from Bacillus subtilis is cloned into Pichia pastoris, and the cloning process is a conventional process;

[0039] (2) Seed preparation stage: the Pichia pastoris obtained after step (1) was cultured on a YPD slant medium to grow a single colony of Pichia pastoris, and then inoculated in a shake flask (100mL YPD medium in a 500mL shake flask) For cultivation, control the rotation speed of the shake flask at 220 rpm, and cultivate in a constant temperature shaker at 30° C. for about 26 hours to obtain shake flask seeds with a wet weight of 56 g / L.

[0040](3) First-level 200L seed tank cultivation: 100LYPD medium is placed in the seed tank, and the shaking flask seeds cultivated in step (1) are inoculated into the first-level seed tank with 2% inoculum,...

Example Embodiment

[0051] Example 2:

[0052] A preparation method of high temperature resistant lipase of the present invention, comprises the following steps:

[0053] (1) Gene cloning process: the high temperature resistant lipase gene derived from Bacillus subtilis is cloned into Pichia pastoris, and the cloning process is a conventional process;

[0054] (2) Seed preparation stage: the Pichia pastoris obtained after step (1) was cultured on a YPD slant medium to grow a single colony of Pichia pastoris, and then inoculated in a shake flask (100mL YPD medium in a 500mL shake flask) For cultivation, control the rotation speed of the shake flask at 200 rpm, and cultivate in a constant temperature shaker at 30° C. for about 31 hours to obtain shake flask seeds with a wet weight of 59 g / L.

[0055] (3) One-level 200L seed tank cultivation: 100LYPD medium is placed in the seed tank, and the shaking flask seeds cultivated in step (1) are inoculated into the first-level seed tank with 2% inoculum, ...

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Abstract

The invention discloses a method for preparing high temperature-resistant lipase. The method comprise the steps of cloning high temperature-resistant lipase genes coming from Bacillus subtilis into pichia pastoris; performing activating treatment on pichia pastoris, and then inoculating pichia pastoris having undergone the activating treatment into a shake flask to perform shake-flask culture; transferring bacterial solution undergoing the shake-flask culture to a seed tank to perform culture; transferring bacterial solution cultured in the seed tank to a fermentation tank to perform fermentation culture; finally adding lipase synergist in fermentation broth, and obtaining the lipase after adsorption and drying. The lipase obtained according to the preparation method has the advantages of resisting high temperature, having a wide pH action sphere and having good tolerance to organic solvents, the applied range of the lipase is expanded, and the problem that the lipase is not resistant to high temperature in feed granulation is solved.

Description

technical field [0001] The invention relates to the field of fermentation, in particular to a preparation method of high temperature resistant lipase. Background technique [0002] Lipase (triacylglycerollipase EC3.1.1.3) is a ubiquitous hydrolase that can catalyze the breaking of triglyceride bonds. Because of its diversity of catalytic functions and its superiority as a biocatalyst, it is widely used in food, tanning, feed, washing, oleochemical and other fields. As a feed additive, lipase has broad application prospects in animal breeding. First, lipase can eliminate lipid anti-nutritional factors in plant-based feed materials, and at the same time, it can make up for the deficiency of lipase in the digestive tract of poultry and young animals. However, in the lipase products that have been industrialized, the pH stability is mostly in the range of neutral and alkaline, and the tolerance to acid is poor, and there are few varieties suitable for feed. [0003] The prepar...

Claims

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Application Information

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IPC IPC(8): C12N9/20C12N15/55C12N15/81
CPCC12N9/20C12N15/81C12N2800/102C12Y301/01003
Inventor 胡爱红朱晓媛莫章桦
Owner HUNAN NANBEIWANG BIOTECH CO LTD
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