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Ketogulonigenium vulgare lacking ED metabolic pathways and 2-KGA production method thereof

A technology of gulonic acid and gulonic acid bacteria, which is applied in the field of microbial fermentation, can solve the problems of unseen natural ketogronic acid bacteria and genetically engineered strains, etc.

Active Publication Date: 2017-03-15
SHENYANG PHARMA UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] In the prior art, there are no reports of natural ketoclonic acid bacteria and genetically engineered strains with the above-mentioned traits

Method used

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  • Ketogulonigenium vulgare lacking ED metabolic pathways and 2-KGA production method thereof
  • Ketogulonigenium vulgare lacking ED metabolic pathways and 2-KGA production method thereof
  • Ketogulonigenium vulgare lacking ED metabolic pathways and 2-KGA production method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1: See SEQ ID: 1, SEQ ID: 2, SEQ ID: 3, and SEQ ID: 4 for the four key enzymes lacking in ketoglobulina SPU B805 and their gene sequences.

Embodiment 2

[0039] Implementation example 2: biological characteristics of ketoglobulina SPU B805

[0040]After Gram staining, SPU B805 is a Gram-negative brevibacterium, which can form short chains or filaments, but cannot form spores. The bacterium was grown on the slant medium for 4 days, and its colony form was point-like, raised, complete, smooth circular colony, and the surface of the colony was oily, transparent, and brown pigment was produced.

[0041] Genomic DNA of strain SPU B805 was extracted, and Primer(S): 5'AGA GTT TGA TCC TGG CTC AG3'; and Primer(A): 5'AAG GAG GTG ATC CAG CCG CA 3' were used as primers for PCR amplification of 16s rRNA After the gene was purified, the gene was sequenced, and the measured sequence was compared with the rRNA homologous sequence in Genbank, and 7 strains with high homology were selected, and the phylogenetic tree of the strain was constructed using the software Mega5. Strain SPU B805 had the highest homology with Ketogulonicigenium vulgare. ...

Embodiment 3

[0042] Implementation example 3: strain SPU B805 shake flask fermentation characteristics

[0043] The mixed bacteria slant (SPU B805 and Bacillus megaterium) was inoculated in the seed medium, and after 18 hours of shaking flask culture at 30°C and 220rpm, it was inoculated in the fermentation medium with 15% replanting amount, and the fermentation medium contained 8% Sorbitose, cultivated at 26-34°C, shaking at 220r / min for 44h. Samples were taken every 4 hours, and the yield and conversion rate of 2-KGA were measured. The experimental results are shown in Table 2. It can be seen that the fermentation of the strain SPU B805 at 32°C for 36 hours can complete the fermentation, and the average conversion rate of 2-KGA can reach 95.34%.

[0044] Table 2. Shake flask fermentation characteristics of strain SPU B805.

[0045]

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Abstract

The invention belongs to the field of microbial fermentation and relates to a bacterial strain for production of 2-keto-L-gulonic acid and a production method thereof, specifically to a bacterial strain for converting a substrate L-sorbose to 2-keto-L-gulonic acid (2-KGA) by the means of microbial fermentation and a technology thereof. By isolating and screening from soil and further artificial domesticating, a new bacterial strain SPU B805, which has stable genetic traits and can match industrial fermentation associated bacterial strain Bacillus megaterium to generate 2-KGA, is obtained. The SPUB 805 is identified as Ketogulonigenium vulgare and has been sent into the China General Microbiological Culture Collection Center (CGMCC) to be preserved. When the SPU B805 matches currently industrial production bacterial strain Bacillus megaterium and under the condition of not changing existing culture technologies, 2-KGA can grow and can be produced at 26-34 DEG C. when concentration of the substrate L-sorbose is 8% and through 32-44 h of fermentation in a shaking flask, conversion rate of 2-KGA is 90% and above. When concentration of the substrate L-sorbose is 10% and through 36-48 h of fermentation in a 5L fermentation tank, conversion rate of 2-KGA can reach 95% and above.

Description

[0001] Technical field: [0002] The invention belongs to the field of microbial fermentation, and relates to a strain producing 2-keto-L-Gulonic acid (2-Keto-L-Gulonic acid, 2-KGA) and a production method thereof, in particular to the use of microbial fermentation to convert a substrate The bacterial strain and process for converting L-sorbose into 2-KGA. [0003] Background technique: [0004] VC is an essential vitamin for the human body. It has a wide range of physiological functions in anti-oxidation and maintaining metabolic balance. It has important uses in the pharmaceutical industry, food industry, feed industry and cosmetics industry. It has a wide range of applications and a huge market. [0005] At present, more than 90% of the world's VC production uses the "two-step fermentation method" invented by my country. The first step of fermentation is to oxidize D-sorbitol to L-sorbose under the action of Gluconobacter oxidans. The second step is to further oxidize L-s...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P39/00C12P7/60C12P17/04C12R1/11C12R1/01
CPCC12P7/60C12P17/04C12P39/00C12N1/205C12R2001/01
Inventor 张怡轩李野
Owner SHENYANG PHARMA UNIVERSITY
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