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Penicillium oxalicum mutant strain a2-13 and its application in preparation of raw amylase preparation and degradation of raw starch

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

Active Publication Date: 2021-10-01
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 its application in preparation of raw amylase preparation and degradation of raw starch
  • Penicillium oxalicum mutant strain a2-13 and its application in preparation of raw amylase preparation and degradation of raw starch
  • Penicillium oxalicum mutant strain a2-13 and its application 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 the Penicillium oxalicum mutant strain A2-13 and its application in preparing raw amylase preparation and degrading raw starch. The present invention provides Penicillium oxalicum A2‑13, whose preservation number in China Center for Type Culture Collection is CCTCC NO: M 2020319. The present invention also provides a method for producing raw amylase or raw amylase preparation with Penicillium oxalicum A2-13, comprising the following steps: cultivating the above-mentioned Penicillium oxalicum A2-13 in a basic or optimized fermentation medium for a certain period of time time, collect the fermentation product, centrifuge and collect the supernatant to obtain raw amylase or raw amylase preparation. Experiments of the present invention prove that the Penicillium oxalicum A2‑13 obtained in the present invention can produce raw amylase preparations, which can efficiently hydrolyze raw tapioca starch and raw corn starch, and the final product of hydrolysis is mainly glucose. This preparation can be used to produce 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 Patents(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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