Method for producing 2,3-butanediol from recombinant Escherichia coli by the aid of glycerin

A technology of Escherichia coli and butanediol, which is applied in the field of recombinant Escherichia coli using glycerol to produce 2,3-butanediol, which can solve problems such as the worrying prospect of sugar as a substrate fermentation production

Inactive Publication Date: 2015-04-29
CHANGZHOU QIUHONG BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
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Problems solved by technology

However, due to the shortage of food, the prospect of fermentation production using sugar as a sub...

Method used

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  • Method for producing 2,3-butanediol from recombinant Escherichia coli by the aid of glycerin

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Embodiment 1

[0111] 1) Cloning of gene fragments

[0112] Using primer sequence 1 and primer sequence 2, the gene fragment alsS was amplified. The PCR amplification conditions were: 95° C. for 30 S, 62° C. for 20 S, 72° C. for 90 S, 30 cycles, and 72° C. for 10 min. Using primer sequence 3 and primer sequence 4, the gene fragment alsD was amplified. The PCR amplification conditions were: 95° C. for 30 S, 62° C. for 20 S, 72° C. for 30 S, 30 cycles, and 72° C. for 10 min. Using primer sequence 5 and primer sequence 6, the gene fragment bdh was amplified, and the PCR amplification conditions were: 95° C. for 30 S, 62° C. for 20 S, 72° C. for 60 S, 30 cycles, and 72° C. for 10 min.

[0113] 2) Construction of plasmid

[0114] The amplified gene fragment alsS was digested with different restriction enzymes KpnI and SphI to obtain sticky ends, and the amplified gene fragment alsD was digested with different restriction enzymes SphI and SalI, To obtain sticky ends, the amplified gene fragment ...

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Abstract

The invention relates to a method for producing 2,3-butanediol from recombinant Escherichia coli by the aid of glycerin. The method comprises steps as follows: 1) cloning genes alsS, alsD and bdh involved in biosynthesis of the 2,3-butanediol; 2) connecting the genes with an Escherichia coli high-copy vector pUC19 to construct a recombinant vector of a coding gene containing protein required by biosynthesis of the 2,3-butanediol; 3) transferring a recombinant plasmid into the Escherichia coli BW25141, and obtaining a strain, which can realize the high yield of the 2,3-butanediol, of the Escherichia coli through selection marking by ampicillin; 4) producing the 2,3-butanediol through fermentation, and recovering the 2,3-butanediol from a fermentation liquid. The method has the advantages of environment-friendliness, high efficiency, stability, low cost and the like.

Description

Technical field: [0001] The invention belongs to the field of microbial genetic engineering. Specifically, three genes involved in the biosynthesis of 2,3-butanediol are cloned and connected to the high-copy vector pUC19 of Escherichia coli, the recombinant plasmid is transferred into Escherichia coli, and ampicillin is used as the Markers are selected to enhance the expression of these three genes in Escherichia coli, so that 2,3-butanediol can be synthesized in a large amount in Escherichia coli. Background technique: [0002] 2,3-Butanediol is an important chemical raw material and liquid fuel, widely used in chemical industry, food, fuel and aerospace and other fields, can be used to produce plastics, prepare antifreeze, and is a good solvent, easy to Conversion to various compounds to produce rubber, flavorings and cosmetics (Syu M J. 2001). It is a colorless, odorless, transparent liquid at room temperature, with a relative molecular weight of 90.12g / mol, and its mole...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12N15/70C12P7/18C12R1/19
Inventor 刘立栋
Owner CHANGZHOU QIUHONG BIOTECH
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