Penicillium oxalicum mutant strain A2-13 and application thereof in preparation of raw amylase preparation and degradation of raw starch

A Penicillium oxalicum and raw starch technology, applied in the biological field, can solve the problems of limited development and utilization, low production of raw amylase, etc.

Active Publication Date: 2021-01-12
GUANGXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the raw amylases of Penicillium and Aspergillus, especially the low yield of raw starch glucoamylase, limit their large-scale development and utilization.

Method used

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  • Penicillium oxalicum mutant strain A2-13 and application thereof in preparation of raw amylase preparation and degradation of raw starch
  • Penicillium oxalicum mutant strain A2-13 and application thereof in preparation of raw amylase preparation and degradation of raw starch
  • Penicillium oxalicum mutant strain A2-13 and application thereof in preparation of raw amylase preparation and degradation of raw starch

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0089] Embodiment 1, the screening of Penicillium oxalicum A2-13

[0090] Penicillium oxalicum A2-13 is obtained from the original fungal strains in the laboratory of the inventor of the present application through multiple rounds of physical and chemical random mutagenesis to obtain a strain with higher amylase activity. Using Penicillium oxalicum (Penicillium oxalicum) genetically engineered strain OXPoxGA15A (CCTCC NO: M 2017794; see Chinese patent application number CN201810816723.7) as the starting strain, three rounds of ethyl methylsulfonate (Ethylmethylsulfone, EMS) mutagenesis were used to obtain A mutant strain E3-16 with improved amylase activity compared with the starting strain OXPoxGA15A. E3-16 is carried out normal pressure and normal temperature plasma (mutagenesis again, obtains a bacterial strain A1-2 with higher amylase activity. Then bacterial strain A1-2 is carried out ARTP and EMS compound mutagenesis and screening, finally obtains mutant strain A2 -13. ...

Embodiment 2

[0119] Embodiment 2, the optimization of amylase condition produced by liquid fermentation of Penicillium oxalicum A2-13

[0120] The experiment was repeated three times, and the specific steps of each repeated experiment were as follows:

[0121] 1. Preparation of spore suspension

[0122] 1. Sterilize the PDA solid medium at 121°C for 30 minutes to make a PDA plate.

[0123] 2. Wash the Penicillium oxalicum A2-13 spores cultivated on the PDA plate for 5 days with 0.1% (v / v) Tween 80 aqueous solution, and the preparation concentration is 1×10 8 per mL of spore suspension.

[0124] 2. Optimization of the initial pH of the medium

[0125] 1. Prepare fermentation media with different pH

[0126] Preparation of basal medium: basal medium consists of wheat bran, crystalline cellulose Avicel (PH-101, Sigma-Aldrich #11365) and saline solution. Wherein, the proportion of wheat bran, crystalline cellulose Avicel and salt solution is 4.0g:1.0g:100mL. Salt solution consists of wat...

Embodiment 3

[0175] Embodiment 3, utilize Penicillium oxalicum A2-13 to prepare raw amylase preparation

[0176] 1. Preparation of fermentation medium

[0177] Prepare the best culture medium for Step 7 of Example 2.

[0178] 2. Preparation of spore solution

[0179] The concentration of preparation embodiment 2 step one is 1 * 10 8 per mL of spore inoculum. Inoculate into the above-mentioned optimum medium at an inoculum amount of 1% (v / v).

[0180] 3. Preparation of raw amylase preparation

[0181] The fermentation medium of the Penicillium oxalicum A2-13 spore suspension inoculated in step 2 was placed in a constant temperature incubator at 28°C for static cultivation for 6 days to obtain a fermentation product; centrifuged (12,000rpm for 15min) to collect the supernatant, the The supernatant is the raw amylase preparation.

[0182] 4. Determination of the raw amylase activity of the raw amylase preparation.

[0183] The results were as follows: the raw amylase activity of the ra...

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Abstract

The invention discloses a Penicillium oxalicum mutant strain A2-13 and application thereof in preparation of a raw amylase preparation and degradation of raw starch. The invention provides the Penicillium oxalicum A2-13, and the preservation number of the Penicillium oxalicum A2-13 in China Center for Type Culture Collection is CCTCC NO: M 2020319. The invention also provides a method for producing raw amylase or the raw amylase preparation by using the Penicillium oxalicum A2-13. The method comprises the following steps of: culturing the Penicillium oxalicum A2-13 in a basic or optimized fermentation culture medium for a certain time, collecting a fermentation product, centrifuging and collecting supernatant to obtain the raw amylase or the raw amylase preparation. Experiments prove thatthe Penicillium oxalicum A2-13 obtained by the invention can be used for producing the raw amylase preparation; the preparation can be used for efficiently hydrolyzing raw cassava starch and raw cornstarch; and in addition, a hydrolysis end product mainly comprises glucose. The preparation can be used for producing alcohol.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a Penicillium oxalicum mutant strain A2-13 and its application in preparing raw amylase preparations and degrading raw starch. Background technique [0002] Amylase is one of the most dominant enzymes in the commercial enzyme market. According to different modes of action, it is mainly divided into α-amylase, glucoamylase, α-glucosidase and starch debranching enzyme. Among them, α-amylase and glucoamylase are the key enzymes in starch processing. α-amylase randomly cuts the α-1,4-glucosidic bonds inside the starch molecular chain to produce a large amount of oligosaccharides and a small amount of maltose and glucose. Glucoamylase can hydrolyze α-1,4-glucosidic bonds and α-1,6-glycosidic bonds, and release glucose from the non-reducing end of the starch molecular chain in turn. A small number of amylase proteins contain a starch-binding domain (Starch-binding dom...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12N9/26C12P19/14C12P19/04C12P19/02C12P19/12C12P7/06C12R1/80
CPCC12N1/14C12N9/2411C12P7/06C12P19/02C12P19/04C12P19/12C12P19/14C12N1/145C12R2001/80Y02E50/10
Inventor 赵帅冯家勋顾丽莎谈明珠
Owner GUANGXI UNIV
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