Engineering bacterium of Geobacillusthermoglucosidasius and construction method and application of engineering bacterium

Abstract

The invention provides an engineering bacterium of Geobacillusthermoglucosidasius and a construction method and application of the engineering bacterium. The construction method includes: through genetic engineering of Geobacillusthermoglucosidasius, introducing ribC (G199D) point mutation, sequentially knocking out purR, purA, ccpN and ldh genes, and then introducing an expression vector carryinga riboflavin synthesis gene cluster into the transformed engineering bacteria to obtain a series of high-temperature riboflavin-producing geobacillus engineering bacteria. The results show that yieldof riboflavin after fermentation of geobacillus after genetic engineering for 24 h is 520 mg / L; and through mixed fermentation with glucose and xylose for the strain, the yield can reach more than 1g / L. The engineering bacterium has advantages of high growth temperature and short generation time, can utilize an inexpensive carbon source, and can be used for large-scale production of riboflavin at low cost.

Description

technical field [0001] The invention relates to the technical fields of genetic engineering and fermentation engineering, in particular, to the thermoglucosidase Geobacillus engineering bacteria and its construction method and application. Background technique [0002] Riboflavin, namely vitamin B2, is one of the essential vitamins for bacteria and animals. It usually exists in the form of FMN and FAD in organisms and participates in intracellular redox reactions. It is mainly used in the fields of pharmaceuticals, feed additives, food additives and cosmetics. At present, the production process of riboflavin is microbial fermentation. In the early stage, its production strains were fungi, such as Candida famata, Ashbya gossypii, Eremothecium ashbyii, Saccharomy cescerevisiae, etc. (Stahmann et al, 2016, Appl Microbiol Biotechnol 100: 2107-2119). However, there are some problems in the production of riboflavin by fungi, such as long fermentation period (6-7 days), low yield...

AI Technical Summary

Problems solved by technology

Second, the production cycle is long, and the growth generation of Bacillus subtilis can reach 40-120min

Third, patent barriers are obvious. Most of the patents on the production of riboflavin by Bacillus subtilis are controlled by large foreign companies, which has caused troubles to the research and application of Bacillus subtilis in China.

Fourth, raw materials are more expensive

Normal temperature bacteria such as Bacillus subtilis produce riboflavin by fermentation at 37°C, and a large number of miscellaneous bacteria can grow at 37°C, which makes it easy to contaminate bacteria during the fermentation process

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