Preparation method and application of Trichoderma reesei strain capable of highly yielding penicillium janczewskii Zaleski alpha-galactosidase

A technology of galactosidase and Penicillium yankii, applied in the biological field, can solve the problems of hidden dangers in biological safety, not reaching the level of industrial production, cumbersome procedures, etc., and achieve simple fermentation process, cost reduction, and cheap and easy-to-obtain raw materials. Effect

Active Publication Date: 2015-12-23
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Abstract
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Problems solved by technology

Among them, the α-galactosidase expressed in Pichia pastoris has the highest yield and good enzyme activity specificity, but it has not yet reached the industrial production level. It needs to be transferred during the fermentation process and needs to be induced with a

Method used

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  • Preparation method and application of Trichoderma reesei strain capable of highly yielding penicillium janczewskii Zaleski alpha-galactosidase
  • Preparation method and application of Trichoderma reesei strain capable of highly yielding penicillium janczewskii Zaleski alpha-galactosidase
  • Preparation method and application of Trichoderma reesei strain capable of highly yielding penicillium janczewskii Zaleski alpha-galactosidase

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Embodiment 1

[0046] Example 1, Preparation of Trichoderma reesei strain with high yield of Penicillium yanchii α-galactosidase

[0047] 1. Preparation of Trichoderma reesei strain with high production of Penicillium yanchii α-galactosidase

[0048] 1. Construction of Penicillium yanchii α-galactosidase expression vector

[0049] (1) Amplification of Penicillium yanchii α-galactosidase gene

[0050] Using the plasmid containing the full sequence of Penicillium yanchii α-galactosidase gene as a template, PCR amplification was carried out using Pagl-F and Pagl-R primers, and the PCR amplification product was obtained, which was the DNA molecule shown in sequence 1 in the sequence listing , which is the gene fragment of Penicillium yanchii α-galactosidase without signal peptide. The primer sequence is as follows (the underline represents the restriction restriction point): Pagl-F: 5'-G GAATTC CAGGACTCAAACGCAAACCCAATCGTG-3'; Pagl-R:5'-G ACTAGT CTAATGATGATGATGATGATGCTGCCTCTCCCAACATCACAA-3'...

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Abstract

The invention discloses a preparation method and application of a Trichoderma reesei strain capable of highly yielding penicillium janczewskii Zaleski alpha-galactosidase. The Trichoderma reesei strain capable of highly yielding the penicillium janczewskii Zaleski alpha-galactosidase is obtained by introducing an encoding gene of the penicillium janczewskii Zaleski alpha-galactosidase and a selection marker gene to a host, namely Trichoderma reesei. The efficient secretory expression of the penicillium janczewskii Zaleski alpha-galactosidase is realized in Trichoderma reesei by utilizing a strong promoter of Trichoderma reesei cbh1 and a signal peptide sequence of the strong promoter. Experiments prove that the enzyme activity of a fermentation solution obtained through fermenting the Trichoderma reesei strain capable of highly yielding the penicillium janczewskii Zaleski alpha-galactosidase can be up to 119.16U/ml; and the fermentation process is simple, the raw materials are cheap and available, the production cost is greatly reduced, and the preparation method is very suitable for industrial production and application.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a preparation method and application of a Trichoderma reesei strain with high yield of Penicillium yanchii α-galactosidase. Background technique [0002] α-galactosidase (α-D-galactosidegalactohydrolase; EC3.2.1.22) is an exoglycosidase that can specifically hydrolyze the α-1,6-galactosidic bond at the non-reducing end of the sugar chain. It can catalyze the hydrolysis of oligosaccharides containing α-galactosidic bonds, such as melibiose, raffinose, stachyose, verbascose, etc., and also catalyze the hydrolysis of heteropolysaccharides containing this bond, such as α-galactomannan sugar etc. Studies have shown that α-galactosidase has great application prospects in industry. For example, in the feed industry, α-galactosidase can effectively remove anti-nutritional factors in soybean meal feed and improve the utilization rate of nutrients in feed; in the sugar in...

Claims

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Application Information

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IPC IPC(8): C12N1/15C12N9/40C12R1/885
Inventor 董志扬林洁陈秀珍马枝枝黄振邦秦丽娜
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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