High-density culture method of clostridium thermocellum

A technology for high-density culture, Clostridium thermocellum, applied in the direction of microorganism-based methods, biochemical equipment and methods, bacteria, etc.

Inactive Publication Date: 2013-01-09
QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] High-density fermentation is the main method to quickly obtain bacteria and their main metabolites, and is also a prerequisite for commercial production. It is of great significance for improving the production intensity and reducing production costs in the fermentation industry (Chen Baoli, Wang Shili, etc., 2009), However, the high-density cultivation of Clostridium thermocellum has not been reported, and the present invention will fill up the gap in this respect

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] Embodiment 1: 100ml anaerobic bottle fermentation

[0017] 1. Use a 1 mL syringe to take 100 μL of C.thermocellum JYT01 strains in glycerol tubes, inoculate them into an anaerobic bottle containing 5 mL of seed medium, and incubate at a constant temperature of 60°C to activate the strains.

[0018] 2. Inoculate 3 mL of the activated C. thermocellum JYT01 bacterial liquid into 50 mL of seed medium, and incubate in a constant temperature incubator at 60°C for 24 hours to obtain a first-grade seed liquid.

[0019] 3. According to the recipe "KH 2 PO 4 1.0g / L, K 2 HPO 4 (anhydrous) 6.0g / L, urea 5.0g / L, MgCl 2 ·6H 2 O 2.49g / L, CaCl 2 2H 2 O 135mg / L, FeSO 4 ·6H 2 O 5.0mg / L, Cysteine ​​hydrochloride 2.0g / L, Resazurin 2.0mg / L, cellobiose 20.0g / L, Morpholinopropane sulfonic acid (MOPS) 10.0g / L, Yeast extract 10.0g / L, Sodium citrate 2H 2 O 0.752g / L" to prepare the fermentation medium, adjust the initial pH to 8.0. Dispense 100mL of the medium into a 300ml anaerobic bot...

Embodiment 2

[0021] Embodiment 2: 5L fermentation tank fermentation

[0022] 1. Prepare 200 ml of C. thermocellum JYT01 primary seed liquid according to the method described in Example 1, and calculate the inoculum size to be 1:10, and the respective required volumes when the total volume of the fermentation broth is 2 L.

[0023] 2. According to the recipe "KH 2 PO 4 1.0g / L, K 2 HPO 4 (anhydrous) 6g / L, urea 17.8g / L, MgCl 2 ·6H 2 O2.49g / L, CaCl 2 2H 2 O 135mg / L, FeSO 4 ·6H 2 O 5mg / L, Cysteine ​​hydrochloride 2.0g / L, Resazurin 2.0mg / L, cellobiose 30.0g / L, Yeast extract 12.0g / L, Sodium citrate 2H 2 O 0.752g / L" to prepare fermentation medium (calcium, magnesium, iron salt mother solution added after sterilization), 115 ℃, 15min moist heat sterilization.

[0024] 3. After the sterilization is completed, nitrogen gas is continuously fed into the fermenter to remove the oxygen in the medium and the fermenter. After the medium changes from light blue or pink (the color of resazurin) t...

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Abstract

The invention belongs to the field of microbial fermentation engineering, and specifically relates to a high-density culture method of clostridium thermocellum. The method is characterized in that a GS-2 culture medium is used as an initial culture medium; the fermentation medium is optimized through a multivariate statistical method; and cell concentration of clostridium thermocellum is rapidly and efficiently raised. The invention has the following advantage: the invention provides a fermentation medium for rapid acquisition of large cell concentration of clostridium thermocellum, thus laying the groundwork for subsequent production of cellulosome which has high conversion efficiency of cellulose to bioethanol. Bacterial density of clostridium thermocellum is raised by 6-9 times in comparison with that of the initial culture medium. OD600 under the bottling condition reaches 8.85, and OD600 in a fermentation cylinder reaches 12.7.

Description

technical field [0001] The invention relates to the field of microbial fermentation engineering, in particular to a method for high-density cultivation of Clostridium thermocellum. Background technique [0002] With the intensification of the global energy and environmental crisis, seeking alternative energy has become a research hotspot in many countries, among which clean and renewable bioenergy is an important aspect of alternative energy research, and bioethanol is the most promising liquid bioenergy One, but bioethanol still mainly comes from sugar and starch fermentation at present, in the face of the rapid expansion of world population and food shortage, the development of producing alcohol with grain will be subject to many restrictions (Wang Dan, Lin Jianqiang, etc., 2002). [0003] Cellulose is the most abundant renewable resource on the earth, but only a small amount of it is effectively used, and most of it is disposed of in the environment as waste, which instea...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12R1/145
Inventor 崔球朱新术张华仁张景涛
Owner QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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