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Method for efficiently producing 1, 3-propylene glycol by reducing by-product acetic acid

A propylene glycol, high-efficiency technology, applied in the field of bioengineering, can solve problems such as no detailed research and no reports, and achieve the effect of fast product synthesis, less by-products, and high conversion rate

Inactive Publication Date: 2015-12-16
EAST CHINA UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The poxB gene is responsible for encoding pyruvate oxidase, which catalyzes the synthesis of acetic acid from pyruvate in organisms; the pta gene is responsible for encoding phosphotransacetylase, which is the key enzyme in catalyzing the synthesis of acetic acid from acetyl-CoA. The specific role in Burgeria has not been studied in detail, and the study of improving the production of 1,3-PD by knocking out these two genes at the same time has not been reported.

Method used

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  • Method for efficiently producing 1, 3-propylene glycol by reducing by-product acetic acid
  • Method for efficiently producing 1, 3-propylene glycol by reducing by-product acetic acid
  • Method for efficiently producing 1, 3-propylene glycol by reducing by-product acetic acid

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1. Knocking out the poxB gene alone severely reduces the growth of bacteria, which is not conducive to the synthesis of 1,3-PD

[0023] The M2014574 strain was cultured overnight at 37° C. in LB medium (0.5% yeast extract, 1% tryptone, 1% NaCl, pH 7.0), and the genome was extracted. Using the extracted M2014574 genome as a template, primers were designed based on the poxB gene (locus_tag: KPN_00904) on the genome sequence of Klebsiella pneumoniae MGH78578 (LOCUS: NC_009648) registered by NCBI, and the target band was recovered after PCR reaction And sequenced, the target band was subjected to gene analysis after sequencing, and the similarity with the poxB gene on MGH78578 was 100%. The poxB gene (1278bp) of the M2014574 genome was knocked out by means of homologous recombination, and the obtained recombinant strain was M2014574ΔpoxB.

[0024] M2014574 and M2014574△pta were inoculated in 250ml Erlenmeyer flasks respectively for anaerobic and aerobic culture at ...

Embodiment 2

[0028] Example 2. Knocking out the pta gene alone has almost no effect on the growth of the bacteria and the synthesis of 1,3-PD

[0029] The M2014574 strain was cultured overnight at 37° C. in LB medium (0.5% yeast extract, 1% tryptone, 1% NaCl, pH 7.0), and the genome was extracted. Using the extracted M2014574 genome as a template, primers were designed based on the pta gene (locus_tag: KPN_02688) on the genome sequence of Klebsiella pneumoniae MGH78578 registered in NCBI (LOCUS: NC_009648), and the target band was recovered after PCR reaction And sequenced, the target band was subjected to gene analysis after sequencing, and the similarity with the poxB gene on MGH78578 was 100%. The poxB gene (2133bp) of the M2014574 genome was knocked out by means of homologous recombination, and the obtained recombinant strain was M2014574Δpta.

[0030] M2014574 and M2014574△pta were inoculated in 250ml Erlenmeyer flasks respectively for anaerobic and aerobic culture at 37°C for 12 hours...

Embodiment 3

[0034] Example 3. Knocking out the poxB and pta genes at the same time did not significantly inhibit the bacteria, but the synthesis of 1,3-PD was improved

[0035] Klebsiella pneumoniae M2014574 was simultaneously knocked out according to the methods in Examples 1 and 2, and the obtained recombinant bacteria were M2014574ΔpoxB-pta.

[0036] M2014574 and M2014574△poxB-pta were respectively inoculated in 250ml Erlenmeyer flasks for anaerobic and aerobic culture at 37°C for 12 hours, and the medium was seed medium supplemented with 40g / L glycerol. During aerobic culture, the flask was sealed with 8 layers of gauze; under anaerobic culture conditions, the air in the flask before inoculation was replaced with nitrogen, and the flask was sealed with a rubber stopper.

[0037] After culturing for 12 hours, the bacterial cell concentration, 1,3-PD and acetic acid concentrations in the fermentation broth were measured, and the results are shown in Table 3. It can be seen from Table 2...

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Abstract

The invention discloses a method for efficiently producing 1, 3-propylene glycol by reducing by-product acetic acid. According to the method, two genes, namely poxB and pta, in klebsiella pneumoniae are knocked out simultaneously, and the 1, 3-propylene glycol is produced by transforming glycerin more efficiently. The method has the advantages that after the two genes are knocked out, synthesis of a by-product, namely acetic acid, is reduced, and the efficiency of producing the 1, 3-propylene glycol by transforming the glycerin with the klebsiella pneumoniae is substantially improved.

Description

technical field [0001] The invention belongs to the technical field of bioengineering. Specifically, two genes related to acetic acid synthesis in Klebsiella pneumoniae are simultaneously knocked out, thereby improving the production level of 1,3-propanediol fermentation. Background technique [0002] 1,3-propylene glycol (1,3-porpanediol, referred to as 1,3-PD) is an important chemical raw material, and its most important use is as a monomer to synthesize a new type of polyester - polytrimethylene terephthalate ( PTT). Studies have shown that PTT is a polyester material with particularly excellent properties, so the industrial value of 1,3-PD has attracted the attention of various countries. Current research shows that among the production methods of 1,3-PD, the bio-legal method has more advantages in industrial application than the chemical synthesis method because of its mild conditions, simple operation, less by-products, and environmental protection. Especially with t...

Claims

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Application Information

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IPC IPC(8): C12P7/18C12N15/74C12R1/22
Inventor 宫衡林杰傅水林
Owner EAST CHINA UNIV OF SCI & TECH
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