Chemostat continuous cultivation device

A cultivation device and chemostat technology, which is applied in the field of continuous chemostat cultivation devices, can solve the problems of hindering the further development and application of technology, long equipment occupation period, easy to be contaminated by operation, etc. Reduced chance and easy operation

Inactive Publication Date: 2012-07-18
NANJING TECH UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Due to the general need for long-term continuous culture, not only has the disadvantages of long equipment occupation period, large power consumption, and high gas consumption, but also has problems such as expensive, special equipment, difficult to apply, and easy to infect bacteria during operation, which seriously hinders the deepening of this technology. development and application

Method used

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  • Chemostat continuous cultivation device
  • Chemostat continuous cultivation device
  • Chemostat continuous cultivation device

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] In this embodiment, Actinobacillus succinogenes (Actinobacillus succinogenes) NJ113, preservation number CGMCC 1716 is used as the starting strain, and the mutant bacteria that are tolerant to high-concentration ammonium ions are continuously cultured and screened in the chemostat of the present invention as an example:

[0043] Medium A (g / L): Glucose 10, K 2 HPO 4 ·3H 2 O 15.5, NaH 2 PO 4 2H 2 O 9.6, NaHCO 3 10, yeast extract 5, corn steep liquor 5, pH 7.2.

[0044] Medium B (g / L): Glucose 20 (growth limiting factor), sodium acetate 1.36, NaCl 1, CaCl 2 0.2, MgCl 2 0.2, Na 2 HPO 4 0.31, NaH 2 PO 4 1.6,K 2 HPO 4 3, yeast paste 10, NH 4 HCO 3 4, 8, 16, 24, 28, C 4 h 4 o 4 Na 2 0,CH 3 COONa·3H 2 O 0, pH 9.0.

[0045] Medium C (solid plate medium) (g / L): glucose 10, K 2 HPO 4 ·3H 2 O 15.5, NaH 2 PO 4 2H 2 O 9.6, NaCl 1.0, corn steep liquor 5, yeast extract 5, NH 4 HCO 3 24. Agar powder 20, bromothymol blue 0.1 (pH 6.0-7.6 yellow to blu...

Embodiment 2

[0053] The implementation steps of the present embodiment are the same as in Example 1, changing the relevant parameter conditions, taking Actinobacillus succinogenes (Actinobacillus succinogenes) CGMCC 1716 as the starting strain, continuous culture and screening in the chemostat of the present invention can utilize inorganic nitrogen Source mutants as an example:

[0054] Medium A: Glucose 20g / L, K 2 HPO 4 ·3H 2 O 31g / L, NaH 2 PO 4 2H 2 O 19.2g / L, NaHCO 3 20g / L, yeast extract 10g / L, corn steep liquor 10g / L, pH 6.5;

[0055] Medium B: glucose 10g / L, sodium acetate 2.72g / L, NaCl 2g / L, CaCl 2 0.4g / L, MgCl 2 0.4g / L, Na 2 HPO 4 0.62g / L, NaH 2 PO 4 3.2g / L, K 2 HPO 4 6g / L, yeast extract 5, 2.5, 0g / L, NH 4 HCO 3 4. 8g / L, C 4 h 4 o 4 Na 2 100,CH 3 COONa·3H 2 O 80, pH 7.0;

[0056] Medium C: Glucose 20g / L, K 2 HPO 4 ·3H 2 O 31g / L, NaH 2 PO 4 2H 2 O 19.2g / L, NaCl 2g / L, corn steep liquor 10g / L, yeast extract 10g / L, NH 4 HCO 3 16g / L, agar powder 20g / ...

Embodiment 3

[0064] In this embodiment, E.coli NZN111 (F-Δpfl::Cam, ldhA::Kan) / pTrc99a-sfcA is used as the starting strain, and the mutant bacteria resistant to organic acids are continuously cultured and screened in the chemostat of the present invention. example:

[0065] Medium A (g / L): Glucose 15, K 2 HPO 4 ·3H 2 O 23.5, NaH 2 PO 4 2H 2 O 14.6, NaHCO 3 15, yeast extract 8, corn steep liquor 7, pH 6.8.

[0066] Medium B (g / L): Glucose 15 (growth limiting factor), sodium acetate 2.13, NaCl 1.5, CaCl 2 0.3, MgCl 2 0.3, Na 2 HPO 4 0.45, NaH 2 PO 4 2.8,K 2 HPO 4 5, yeast paste 5, NH 4 HCO 3 8,C 4 h 4 o 4 Na 2 200,CH 3 COONa·3H 2 O 180, pH 8.0.

[0067] Medium C (solid plate medium) (g / L): glucose 15, K 2 HPO 4 ·3H 2 O 23.2, NaH 2 PO 4 2H 2 O14.6, NaCl 1.5, corn steep liquor 7, yeast extract 8, NH 4 HCO 3 20, agar powder 20, bromothymol blue 0.15 (pH 6.0-7.6 yellow to blue), pH 7.1.

[0068] Using the device of the present invention, continuously cult...

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Abstract

The invention relates to a chemostat continuous cultivation device which is characterized by comprising an acid-alkali pH adjusting and controlling system, a supplementing system, a reaction system, a discharging and sampling system and a gas inlet system, wherein the reaction system comprises a reaction tank which is respectively communicated with the acid-alkali pH adjusting and controlling system, the supplementing system, the discharging and sampling system and the gas inlet system through connecting pipes, and a sealing member is arranged at a bottle opening of the reaction tank. The invention also relates to a method for screening a succinic acid mutant strain by using the chemostat continuous cultivation device of the invention. The device of the invention has simple materials, needs no additional special equipment, no stirring device, and features easy assembly, convenient use, small equipment, little power consumption, little gas consumption, good sealing property and difficult microbiological contamination; and the method of the invention not only features simple operation, durability and stability, but also can realize continuous cultivation on a long-term basis.

Description

technical field [0001] The invention belongs to the field of industrial biochemical industry, and in particular relates to a chemostat continuous culture device, and also relates to a method for screening microorganisms using the device. Background technique [0002] As an important C4 platform compound, succinic acid (also known as succinic acid) is widely used in the fields of medicine, food industry, agricultural production and chemical industry. The intermediate chemicals based on succinic acid currently in the research and development stage mainly include: 1,4-butanediol (BDO), γ-butyrolactone, tetrahydrofuran, N-methylpyrrolidone (NMP), hexamethylene diol Acid, malic acid and new biodegradable plastic product polybutylene succinate (PBS). The traditional production method uses butane to be produced by electrolysis through maleic anhydride, which causes great pollution and high cost, which seriously inhibits the development potential of succinic acid. [0003] Biologi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12M1/36C12M1/00C12N1/20C12Q1/02
CPCC12M41/26
Inventor 姜岷叶贵子陈可泉李建马江峰韦萍欧阳平凯
Owner NANJING TECH UNIV
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