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Method for preparing and regenerating aspergillus oryzae protoplast for high-yield lactase

A protoplast and Aspergillus oryzae technology, which is applied in the field of preparation and regeneration of Aspergillus oryzae protoplasts, can solve the problem of limited expression and achieve high regeneration rate and high number of preparations

Inactive Publication Date: 2017-11-21
QILU UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although some progress has been made, the amount of expression is still limited

Method used

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Embodiment 1: a kind of preparation and the regeneration method of the aspergillus oryzae protoplast that is used for high-yielding lactase comprises the following steps:

[0038] (1) Activation culture of strains:

[0039] Fully dissolve the freeze-dried and preserved Aspergillus oryzae strain (BNCC338380) into a bacterial suspension with sterile physiological saline, inoculate the bacterial suspension on the slant of Chapeauer's agar medium for activation, and cultivate it at 28°C for 6 days until it is covered with yellow-green After the spores are used for later use, the continuous expansion is carried out once.

[0040] The culture medium used during the expansion is Chapei's medium.

[0041] (2) Preparation of single spore suspension

[0042] After eluting the yellow-green spores covered with Cha's culture medium with sterile physiological saline, pour it into a 250ml sterile Erlenmeyer flask equipped with glass beads, fully shake the Erlenmeyer flask, under the...

Embodiment 2

[0062] Embodiment 2: a kind of preparation and the regeneration method of the Aspergillus oryzae protoplast for high-yielding lactase comprise the following steps:

[0063] (1) Activation culture of strains:

[0064] Fully dissolve the freeze-dried and preserved Aspergillus oryzae strain (BNCC338380) into a bacterial suspension with sterile physiological saline, inoculate the bacterial suspension on the slant of Chapeauer's agar medium for activation, and cultivate it at 28°C for 5 days until it is covered with yellow-green After the spores are used for later use, the continuous expansion is carried out once.

[0065] The culture medium used during the expansion is Chapei's medium.

[0066] (2) Preparation of single spore suspension

[0067] After eluting the yellow-green spores covered with Cha's culture medium with sterile physiological saline, pour it into a 250ml sterile Erlenmeyer flask equipped with glass beads, fully shake the Erlenmeyer flask, under the action of gla...

Embodiment 3

[0087] Embodiment 3: a kind of preparation method for the aspergillus oryzae protoplast of high-yield lactase comprises the following steps:

[0088] (1) Activation culture of strains:

[0089] Fully dissolve the freeze-dried and preserved Aspergillus oryzae strain (BNCC338380) into a bacterial suspension with sterile physiological saline, inoculate the bacterial suspension on the slant of Chapeauer's agar medium for activation, and cultivate it at 28°C for 5 days until it is covered with yellow-green After the spores are used for later use, the continuous expansion is carried out once.

[0090] The culture medium used during the expansion is Chapei's medium.

[0091] (2) Preparation of single spore suspension

[0092] After eluting the yellow-green spores covered with Cha's culture medium with sterile physiological saline, pour it into a 250ml sterile Erlenmeyer flask equipped with glass beads, fully shake the Erlenmeyer flask, under the action of glass beads, Disperse the...

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Abstract

The invention relates to a method for preparing and regenerating aspergillus oryzae protoplast for high-yield lactase. The method comprises the following steps: (1) preparing aspergillus oryzae spore suspension; (2) performing physiochemical mutation so as to obtain a mutant strain; (3) performing enzymolysis to prepare protoplast; (4) purifying the protoplast; (5) regenerating the protoplast. The quantity of the protoplast prepared by the method disclosed by the invention is high, the obtained protoplast has high activity, and the regeneration rate can reach 31.2%. The protoplast is high in preparation rate and excellent in activity maintenance, is favorable for preparation of subsequent fusants and has significances for researching the metabolic regulation theory of aspergillus oryzae regulated lactase and improving the lactase activity; meanwhile, a novel pathway is provided for strain breeding during lactase production.

Description

technical field [0001] The invention relates to the biological field of fungi, in particular to a method for preparing and regenerating protoplasts of Aspergillus oryzae with high lactase production. Background technique [0002] Lactase, the system name is β-D-galactoside galactohydrolase, or β-galactosidase for short. Lactase has a wide range of applications in food industry, pharmaceutical industry, environmental protection, analysis and so on. Utilizing the ability of lactase to hydrolyze lactose to produce low-lactose products (such as lactose hydrolyzed milk, low-lactose milk powder, etc.), can effectively solve the problem of "lactose intolerance", which is one of the largest uses of lactase in the food industry. [0003] Compared with foreign countries, the consumption of dairy products in my country is very different. The reason for this phenomenon is not only the level of economic development, but also the fact that a large part of the Chinese population suffers f...

Claims

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Application Information

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IPC IPC(8): C12N1/14C12N15/01C12N9/24C12R1/69
CPCC12N1/14C12N9/2402C12N15/01C12Y302/01108C12R2001/69C12N1/145
Inventor 孙华于功明秦大伟
Owner QILU UNIV OF TECH
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