Recombinant pichia pastoris engineering bacteria and production method thereof

Abstract

The invention discloses recombinant pichia pastoris engineering bacteria. The recombinant pichia pastoris engineering bacteria comprises a codon-optimized Richter trichoderma xylanase gene, and the nucleotide sequence of the gene is shown in SEQ:1 in a sequence table. The invention also discloses a production method of the recombinant pichia pastoris engineering bacteria. The production method comprises the following steps: (1) performing synonymous mutation on the Richter trichoderma xylanase gene according to pichia pastoris flavored codons so that the Richter trichoderma xylanase gene is completely mutated into the pichia pastoris favored codons; (2) synthesizing an optimized xylanase gene by utilizing a total-gene synthesis technology, and connecting the optimized xylanase gene with a pMD20-T vector to build a cloning vector pMD20T-Xynopti; (3) building a recombinant expression vector pPICZalpha-Xynopti; (4) transforming a pichia pastoris host; (5) performing induction culture of a positive converter; (6) producing xylanase by small-scale fermentation by adopting the recombinant pichia pastoris engineering bacteria. The recombinant pichia pastoris engineering bacteria disclosed by the invention can be used for secreting a great amount of xylanase.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a recombinant Pichia pastoris engineering bacterium and a production method thereof. Background technique [0002] Hemicellulose is the second most abundant available natural resource after cellulose content. Xylan is an important component of hemicellulose in the cell wall of plant materials, exists under lignin and cellulose, and is tightly combined with lignin and cellulose. Xylan is a kind of poly five-carbon sugar, which is mainly connected by multiple same or different five-carbon sugars through 1,4-β-D-xylosidic bonds. It is one of the main anti-nutritional factors in feed. Xylanase (xylanase) is a kind of glycoside hydrolase, which can catalyze the endohydrolysis of 1,4-β-D-xylosidic bonds in xylan molecules, and its main hydrolysis products are xylobiose and xylobiose Xylo-oligosaccharides, and a small amount of xylose and arabinose, this effect not only reduces the ...

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Problems solved by technology

[0005] Although the application fields of xylanase are very wide, but due to the low yield of xylanase in natural strains, high production cost, low activity of isolated glycanase, and poor enzyme production efficiency, the enzymatic properties have not reached industrial application Requirements, these all restrict the application and development of xylanase

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