Inocula producing cellulase and use thereof

A technology of cellulase and bacterial agent, applied in the direction of application, enzymes, enzymes, etc., can solve the problems of unguaranteed expression and yield, long fermentation cycle, high equipment and floor space

Inactive Publication Date: 2009-06-24
北京挑战生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The former method needs to produce three kinds of enzymes separately, the whole fermentation period is long, and the requirements for equipment and floor space are relatively high.
However, the second method express

Method used

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  • Inocula producing cellulase and use thereof
  • Inocula producing cellulase and use thereof
  • Inocula producing cellulase and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0039] Example 1. Bacteria for producing cellulase

[0040] 1. Cloning of endoglucanase gene

[0041] Bacillus amyloliquefaciens CICC 20467 (China Industrial Microbial Culture Collection and Management Center) was cultured in LB medium (0.5g / 100ml yeast extract, 1.0g / 100ml peptone, 1.0g / 100ml NaCl, pH7.0) for 12h to obtain The fresh bacterial liquid of Bacillus amyloliquefaciens is used for PCR amplification of endoglucanase gene. The nucleotide sequence of the primers used is as follows:

[0042] 5’ primer: GAATTC GCAGGGACAAAAACGCCA (underlined is EcoRI recognition site);

[0043] 3’ primer: GCGGCCGC CTA ATTTGGTTCTGTTCCCC (underline is Not I recognition site, italic is stop codon).

[0044] The PCR reaction system is as follows:

[0045]

[0046]

[0047] PCR amplification conditions: 94°C 3min; 94°C 30s, 53.5°C 30s, 72°C 1min30s, a total of 28 cycles; 72°C 10min.

[0048] A fragment of 1413 bp was obtained by PCR amplification, and the fragment was cloned into the pEASY-T3 ...

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Abstract

The invention discloses a microbial inoculum of cellulase production and the application thereof. The active ingredients of the microbial inoculum of the cellulase production comprise pichia stipitis 1413, pichia stipitis 1515 and pichia stipitis 2526; wherein, the pichia stipitis 1413 is recombinant bacterium obtained by introducing endoglucanase gene into the pichia stipitis, the pichia stipitis 1515 is recombinant bacterium obtained by introducing exoglucanase gene into the pichia stipitis, and the pichia stipitis 2526 is recombinant bacterium obtained by introducing beta-glucosidase gene into the pichia stipitis. The cellulose can be produced by fermenting the microbial inoculum of cellulase production.

Description

technical field [0001] The invention relates to a cellulase-producing bacterial agent and its application. Background technique [0002] The traditional cellulase production method is mainly to screen excellent enzyme-producing strains in nature, and then obtain enzyme-producing strains for industrialized large-scale production through certain mutagenesis methods. However, the degradation of cellulose in the natural environment is accomplished by a variety of microorganisms. These microorganisms coexist with each other to form a micro-ecological system that degrades cellulose. In a specific ecosystem, microorganisms cooperate with each other and restrict each other, so that the ratio of the enzymes that produce cellulose is in an optimal state for degrading cellulose, and once the microorganisms that produce cellulase from If it is separated from its niche, it will destroy this optimal ratio. When screening excellent strains, it is undoubtedly separated from its ecological...

Claims

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Application Information

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IPC IPC(8): C12N1/19C12N9/46C12N9/42C12N15/56C12R1/84
Inventor 吴培均李卓夫李富伟段文娟
Owner 北京挑战生物技术有限公司
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