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Bacillus subtilis and method for fermentation production of D- ribose

A technology of Bacillus subtilis and ribose, applied in the field of microbial fermentation, can solve the problems of high residual sugar, easy to produce spores, long fermentation cycle, etc.

Active Publication Date: 2013-06-12
SHANDONG JINYANG PHARMA +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The object of the present invention is to provide a kind of Bacillus subtilis that loses spore forming ability, stable high-yield D-ribose, and the method for producing D-ribose by fermentation of the strain, to overcome the high residual sugar in the prior art, easy to produce spores, and fermentation cycle long defect

Method used

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  • Bacillus subtilis and method for fermentation production of D- ribose

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Embodiment 1

[0040] Cultivate Bacillus subtilis ATCC21360 on a slant medium at 35-38°C for 2 days, scrape the bacterial cells into sterile saline, and make a bacterial suspension with a cell concentration of 1×10 8 ~10 9 individual / mL. According to conventional mutagenesis methods, physical and chemical mutagens such as ultraviolet lamps and nitrosoguanidine are used for alternate phase or continuous treatment. Take the above-mentioned bacterial suspension in a Petri dish, place it under a 15W ultraviolet lamp at a distance of 20cm, and irradiate it for 0.5-5 minutes; take the above-mentioned bacterial suspension and treat it with 1 mg / mL nitrosoguanidine solution, and incubate and shake at 35-38°C for 30 minutes. For ~60 minutes, centrifuge the cells treated with nitrosoguanidine and wash them with normal saline several times to remove residual nitrosoguanidine. Spread the bacterial solution treated with ultraviolet light, nitrosoguanidine alternately or continuously on the plate medium...

Embodiment 2

[0049] The Bacillus subtilis transketolase-deficient mutant strain BAV20120416 obtained by mutagenesis was inoculated on a slant medium, and cultured at 36° C. for 2 days to obtain a slant strain. Take a ring of slant bacteria and inoculate it into a 500mL Erlenmeyer flask containing 50mL of seed medium, culture it on a shaker at 36°C and 150 r / min for 14 hours, and insert 10% of the inoculum into the pre-prepared 500mL fermentation medium In a 5L shake flask, at 36°C, with a speed of 150r / min, the shaker fermentation culture was carried out. After 48 hours of fermentation, the glucose content in the fermentation broth dropped to 0, and the D-ribose content reached 90.2g / L; while its parent strain Under the same conditions, ATCC21360 needs to be cultured for 72 hours, and the glucose content in the fermentation broth will not drop after it drops to 4 g / L, and the D-ribose content in the fermentation broth is only 29.7g / L.

[0050] The above slant medium composition (%): pepton...

Embodiment 3

[0054] The Bacillus subtilis transketolase-deficient mutant strain BAV20120416 obtained by the above mutagenesis was inoculated on the slant medium of eggplant bottles, and cultured at 37° C. for 2 days to obtain a large slant strain. Add sterile water to the eggplant bottle to wash the strains to make a bacterial suspension, inoculate the bacterial suspension into a 5L shaker flask with 800mL of seed medium, and culture it on a shaker at 37°C with a speed of 150r / min for 16 hours , put 10% of the inoculum into the pre-prepared fermentation medium, the 50L fermentation tank has a liquid volume of 30L, and carry out ventilation and stirring culture at 37°C, wherein the stirring speed is 200~300r / min, and the ventilation volume is 1~2Nm 3 / h, the pressure in the tank is 0.05Mpa, the initial pH of the fermentation broth is 6.5~7.2, the pH of the fermentation broth is controlled at 5.4~7.0 by adding acid and alkali during the fermentation process, and the glucose and D-ribose in th...

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Abstract

The invention discloses a bacillus subtilis BAV20120416 bacterial strain and a method for fermentation production of a D- ribose by the bacterial strain. The bacterial strain is preserved in China General Microbiological Culture Collection Center (CGMCC) on April, 16th in 2012. The preservation number of the bacterial strain is CGMCC NO.6013. The bacterial strain is a bacillus subtilis transketolase deletion mutation strain, has the advantages that hereditary stability is good, spores are difficult to generate and the like, and is capable of accumulating more than 90g / L of the D-ribose in a fermentation medium with glucose and corn steep liquor as main raw materials. The D-ribose accumulated by the bacterial strain is about third more than that accumulated by a parent strain of the bacterial strain, wherein the parent strain is bacillus subtilis ATCC21360. The fermentation time is short and generally controlled to be about 50 hours which is one-third lower than the time in the prior art. Particularly, impurities are few. Residual sugar of final fermentation liquid is reduced to 0. The bacterial strain is particularly applicable to the requirements for medicine production.

Description

technical field [0001] The invention belongs to the technical field of microbial fermentation, and in particular relates to a bacillus subtilis and a method for producing D-ribose by fermentation of the bacterium. Background technique [0002] D-ribose is a very important aldopentose, an important component of genetic material nucleic acid in organisms, and an important intermediate metabolite for organisms to synthesize some amino acids, bases, etc., and has important physiological functions. [0003] D-ribose was first used in semi-synthetic production of vitamin B 2 . With the development of the D-ribose industry and the in-depth development of its application research, its prospects in the food industry and the pharmaceutical industry have gradually been recognized. D-ribose can be used as a raw material for the synthesis of nucleotide flavor enhancers, such as 5'-IMP, 5'-GMP, etc.; in the pharmaceutical industry, its research and application as a raw material for synt...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P19/02C12R1/125
Inventor 蔡会英
Owner SHANDONG JINYANG PHARMA
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