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Colibacillus capable of generating succinic acid by utilizing synthetic medium pure anaerobic growth and application thereof

A synthetic medium and a technology for producing succinic acid, applied in the direction of microorganism-based methods, bacteria, microorganisms, etc., to achieve significant social significance and economic value

Inactive Publication Date: 2012-08-22
NANJING UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, under pure anaerobic conditions, there are few reports on the growth and accumulation of succinic acid in Escherichia coli using synthetic media.

Method used

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  • Colibacillus capable of generating succinic acid by utilizing synthetic medium pure anaerobic growth and application thereof
  • Colibacillus capable of generating succinic acid by utilizing synthetic medium pure anaerobic growth and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] This example illustrates the Escherichia coli ( Escherichia coli ) BER108 screening method.

[0028] 1. The method for the first step plasma mutagenesis of the original strain of Escherichia coli is as follows:

[0029] coli E. coli The original strain of AFP111 was activated in LB test tubes, cultured overnight at 37°C, 200 r / min; freshly cultured cells were diluted to cell concentration OD 600 =1.0, drop it on a sterile slide, dry it with sterile wind; use helium as the discharge gas, 80~120W as the radio frequency power, 10~30SLM as the gas flow rate, and 10~30s as the irradiation time, Plasma mutagenesis was performed on the strain; after mutagenesis, the bacterial film on the slide was eluted with 1 mL of sterile saline, spread on a synthetic medium plate, and cultivated in an anaerobic box.

[0030] In the above screening method, in the plasma mutagenesis method, the radio frequency power is preferably 100W, the gas flow rate is 20SLM, and the irradiation tim...

Embodiment 2

[0046] This example demonstrates that Escherichia coli ( Escherichia coli ) Passage stability of BER108.

[0047] On the synthetic medium plate, the mutant strain BER108 was transferred and cultured several times, and the obtained strains were subjected to fermentation verification respectively. The experimental results are shown in Table 2:

[0048]

[0049] From the experimental results, it can be seen that after 8 consecutive subcultures, the succinic acid production and succinic acid conversion rate of the mutant strain BER108 are relatively stable, which has good passage stability and can be used as a production strain for further research and development.

Embodiment 3

[0051] This example demonstrates that Escherichia coli ( Escherichia coli ) BER108 fermentation process for the production of succinic acid.

[0052] The culture medium formula described in the present embodiment:

[0053] Plate medium: citric acid 3 g L -1 , Na 2 HPO 4 12H 2 O 4 g L -1 , KH 2 PO 4 8 g L -1 , (NH 4) 2 HPO 4 8 g L -1 , NH 4 Cl 0.2 g L -1 , (NH 4 ) 2 SO 4 0.75 g·L -1 , MgSO 4 ·7H 2 O 1 g L -1 , CaCl 2 2H 2 O 10.0 mg·L -1 , ZnSO 4 ·7H 2 O 0.5 mg L -1 , CuCl 2 2H 2 O 0.25 mg L -1 , MnSO 4 ·H 2 O 2.5 mg L -1 , CoCl 2 ·6H 2 O 1.75 mg L -1 , H 3 BO 3 0.12 mg·L -1 , Al 2 (SO4 ) 3 1.77 mg·L -1 , Na 2 MoO 4 2H 2 O 0.5 mg L -1 , ferric citrate 16.1 mg·L -1 , agar 20g / L, glucose 10g / L.

[0054] Seed medium: peptone 10g / L, yeast powder 5g / L, NaCl 5g / L.

[0055] Fermentation medium: citric acid 3 g L -1 , Na 2 HPO 4 12H 2 O 4 g L -1 , KH 2 PO 4 8 g L -1 , (NH 4 ) 2 HPO 4 8 g L -1 , NH 4 Cl 0.2 g L -1 , (N...

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Abstract

The invention relates to a colibacillus capable of generating succinic acid by utilizing synthetic medium pure anaerobic growth. The colibacillus is classified and named as EscherichiacoliBER108 and has the preservation number of CCTCC (China Center for Type Culture Collection) NO.M2012068. The invention further discloses an application of the colibacillus to fermentation production of succinic acid. According to the invention, plasma is utilized to induce colibacillus, a synthetic medium flat screen is utilized to screen out a strain capable of growing rapidly under an anaerobic condition, and under the anaerobic condition, the strain grows by utilizing an inorganic nitrogen source and glucose and accumulates succinic acid; the strain is fermented in a shake flask for 72 hours by utilizing basic magnesium carbonate as a pH regulator; the OD600 of the strain reaches 7.6, the succinic acid yield reaches 11.2g / L, the yield of the succinic acid is increased by nearly three times compared with that of an original strain; and the original strain grows slowly and is low in acid yield under the pure anaerobic synthetic medium condition, so that the mutant strain BER108 has great social significance and economic value.

Description

technical field [0001] The invention belongs to the technical field of industrial microorganism breeding and its fermentation, and relates to an Escherichia coli that uses a synthetic medium for pure anaerobic growth and produces succinic acid. Background technique [0002] Succinic acid, also known as succinic acid, is a common natural organic acid widely found in the human body, animals, plants and microorganisms. As an intermediate product of the TCA cycle and one of the terminal reduction products of anaerobic metabolism, succinic acid plays a very important role in the process of biological metabolism. Research results in recent years have shown that succinic acid can also be used as a C4 platform compound to synthesize bulk chemicals such as 1,4-butanediol, tetrahydrofuran, and γ-butyrolactone, as well as polybutylene succinate (PBS) biodegradable compounds. Degrades polyester. In recent years, with the increasing depletion of fossil resources and the increasingly se...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P7/46C12R1/19
Inventor 姜岷张常青苟冬梅梅佳军刘嵘明马江锋吴明科曹伟佳
Owner NANJING UNIV OF TECH
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